Topical, isotonic compositions for genital use

ABSTRACT

The present disclosure provides compositions and methods for using topical, isotonic compositions comprising a prebiotic oligosaccharide, a metal co-factor, and an essential oil comprising bornyl acetate. The compositions support the genital microbiota and are useful for, for example, hydrating, lubricating, cleaning, and/or decreasing irritation or inflammation of the urogenital and/or anogenital region of a subject, and/or enhancing the beneficial genital microbiota of a subject. Such compositions are useful before, during, and/or after sexual and/or reproductive activity.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation application relating to and claimingthe benefit of commonly-owned, co-pending PCT International ApplicationNo. PCT/US2019/065548, filed Dec. 10, 2019, which claims priority to andthe benefit of U.S. Provisional Patent Application. No. 62/778,278,filed Dec. 11, 2018, the contents of each of the foregoing are herebyincorporated by reference in its entirety.

BACKGROUND

The genital microbiome of a human is a unique combination of microbialspecies comprising at least one hundred species of bacteria and avariety of fungal, viral, and protozoal species. There is considerablevariation in make-up of the genital microbiome between individuals, withmany factors such as hygiene regimes, diet, environment, age, ethnicity,disease, sexual activity, sexual orientation, and life history affectingthe presence of specific microbial species and their metabolicactivities. Environmental conditions within the genitalia compared toother locations of the human body are distinct. As a result, productsindicated for urogenital, anogenital, vaginal and/or penile use, tomodulate the genital microbiome, may require distinct prebiotic and/orprobiotic constituents.

Additionally, the genital microbiome is unique in that it is sharedbetween members of a sexual dyad and between mammalian mother andnewborn as a primary mode of the newborn's establishment of its ownmicrobiome. Diseases and/or dysbiosis within the dyad can occur if ahealthy genital microbiome is not supported. Microbiome congruency andtransfer of microbial species between sexual partners, and between amother and her child is well documented. Sexual partners can transmitpathogenic bacteria back and forth that harm beneficial bacteria,increase inflammation, and increase risk for disease, includinginfertility, reproductive dysfunction, poor pregnancy outcomes,autoimmune disease, sexually transmitted diseases (e.g., HIV, HSV, HPV),cancers (e.g., prostate, cervical), and systemic diseases such ascognitive impairment.

Furthermore, elderly and children/infants often have disrupted genitalmicrobiomes, especially due to wearing of barrier protection diapers andchronic incontinence, which can lead to chronic inflammation anddisease. Improving the healthy bacteria in the urogenital and/oranogenital region during cleansing can improve their overall health.Continuous cleaning of the urogenital and/or anogenital region usingcurrent soaps or cleansing wipes can irritate the skin and disrupt themicrobiome. Continuous handwashing by care providers with products thatchange skin pH and harm beneficial bacteria can limit healthy microbiomeof the caregiver, decreasing the opportunity for transfer of beneficialbacteria to the seniors or children to help repopulate a healthymicrobiome.

Existing cleaning products and genital therapies (e.g., body washes,wipes, yeast treatments, most lubricants) have pH, salt levels andingredients that harm genital tissues and kill healthy bacteria. Forexample, a popular, commercially available diaper wipe has a pH of 3.While the World Health Organization (WHO) has state that this pH levelis inconsistent with human genital tissues. The WHO provided anEmergency Advisory asking for products in contact with the genital andrectal region to have safe salt (ion), pH and ingredient formulas, asdisclosed in a World Health Organization Advisory Note “Use andprocurement of additional lubricants with male and female condoms:WHO/UNFPA/FHI360” (2012), hereby incorporated by reference in itsentirety, and particularly in relation to the pH and Osmolality ofcommercial lubricants in Annex 1. However, commercial products have notchanged.

There remains a need for compositions for use in the urogenital and/oranogenital region that are not harmful to the genital tissues and thatpreserve and support the beneficial regional microbiome of these areas.Presently disclosed embodiments address this need and provide otherrelated advantages.

SUMMARY

The present disclosure provides compositions and methods for maintainingthe microbiome of certain subject bodily regions including theurogenital (e.g., subject regions relating to function of urinaryexcretion and reproduction) and/or anogenital regions (e.g., relating tothe anus and genitals). In particular, the present disclosure relates toisotonic, genital microbiome-friendly topical compositions comprising aprebiotic oligosaccharide, a metal co-factor, and bornyl acetate (e.g.,via an essential oil comprising bornyl acetate), at a pH level tocomplement a subject's gender, life-stage and use. The isotonic, genitalmicrobiome-friendly compositions of the present disclosure can be usedfor hydrating the urogenital and/or anogenital region of a subject,lubricating the urogenital and/or anogenital region of a subject,cleaning the urogenital and/or anogenital region of a subject,decreasing irritation or inflammation of the urogenital and/oranogenital region of a subject, enhancing the genital microbiota of asubject (e.g., increasing the colony count of beneficial genitalmicrobiota of a subject), enhancing genital tissue function of asubject, or the like.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 depicts an applicator of the present disclosure which may be usedfor delivery of compositions to the urogenital and/or anogenital region(e.g., vagina).

FIG. 2 depicts the use of the applicator shown in FIG. 1.

FIG. 3A depicts an applicator of the present disclosure in storageconfiguration. FIG. 3B depicts the applicator in FIG. 3A nearly in theapplication configuration. FIG. 3C depicts the applicator in thedisposal configuration.

FIG. 4A depicts an applicator of the present disclosure disassembled(e.g., as it may come in a kit). FIG. 4B depicts the applicator of FIG.4A being assembled.

FIG. 5 shows the average pH values of various regions of the penis (n=6)with a forearm (ante fossa) control. Error bars illustrate the standarddeviation.

FIG. 6 shows the average pH values of various regions of the penis witha forearm (ante fossa) control following administration of a foamingwash composition of the disclosure (Formulation A) and a conditioner ofthe disclosure (Formulation B). For comparison, pH values are alsoillustrated for a commercial Nivea wash followed by Astroglideconditioner. Error bars illustrate the standard deviation for eachmeasurement.

FIG. 7A shows the percent total motility of sperm as compared to controlmeasured in the bovine sperm motility studies. FIG. 7B shows the percentof progressively motile sperm compared to control in the bovine spermmotility studies. Formulations tested were 71519A, 71519B, 71519C,0618CA45, and 0618CA68 (Table 15).

FIG. 8A shows the percent total motility of sperm with pH of 6.8formulation testing as compared to control. FIG. 8B shows the percent ofprogressive motility of sperm with pH of 6.8 formulation testing.Formulations tested were 060919C, 0617HF, 0617HL, and 0617HM (Table 16).

FIG. 9 shows the mucoadhesion of porcine mucus admixed with a lubricantcomposition of the disclosure (Table 21) as compared to commercialRephresh and Replens lubricants. Error bars represent the standarddeviation.

DETAILED DESCRIPTION

Prior to setting forth this disclosure in more detail, it may be helpfulto an understanding thereof to provide definitions of certain terms tobe used herein. Additional definitions are set forth throughout thisdisclosure.

In the present description, any concentration range, percentage range,ratio range, or integer range is to be understood to include the valueof any integer within the recited range and, when appropriate, fractionsthereof (such as one tenth and one hundredth of an integer), unlessotherwise indicated. Also, any number range recited herein relating toany physical feature, such as polymer subunits, size or thickness, areto be understood to include any integer within the recited range, unlessotherwise indicated. Any concentration ranges recited herein are to beunderstood to include concentrations of any integer within the range andfractions thereof, such as one tenth, one hundredth, and one thousandthof an integer, unless otherwise indicated. Unless otherwise indicated,it will be understood that any percentage refers to the weightpercentage with respect to the indicated component. Typically, thepercent of a component in the composition indicates the weightpercentage with respect to the weight of the composition. As usedherein, the term “about” means±20% of the indicated range, value, orstructure, unless otherwise indicated. The term “consisting essentiallyof” is not equivalent to “comprising” and refers to the specifiedmaterials or steps, or to those that do not materially affect the basicand novel characteristics of the claimed subject matter. It should beunderstood that the terms “a” and “an” as used herein refer to “one ormore” of the enumerated components. The use of the alternative (e.g.,“or”) should be understood to mean either one, both, or any combinationthereof of the alternatives. As used herein, the terms “include,” “have”and “comprise” are used synonymously, which terms and variants thereofare intended to be construed as non-limiting.

As used herein, the term “dyad” refers to a group of two persons havinga sociologically significant relationship (e.g., sexual relationship,parent-infant relationship, parent-child relationship, health careprovider and patient relationship, care-giver and patient relationship).A dyad may be composed of two males, two females, one male and onefemale, or other gender groupings (e.g. non-binary gendered individual,intersex individual). In certain embodiments, a dyad is a sexual dyad.In certain embodiments, a dyad is a mother-fetus, parent-infant orparent-child dyad. In certain embodiments, a dyad is a caregiver-patientdyad.

The term “non-monadic sexual relationship” refers to a group of two ormore persons having a sociologically significant sexual relationship. Incertain embodiments, a non-monadic sexual relationship is a sexual dyad.A sexual dyad may be a heterosexual dyad, a homosexual dyad, or othersexual orientation dyad. In other embodiments, a non-monadic sexualrelationship is a sexual triad (a group of three people).

As used herein, the term “genital microbiota” or “genital microbiome”refers to the collective microorganisms that normally colonize thegenital region and are non-pathogenic. The genital microbiota may referto that of the genital skin microbiota, the vaginal microbiota (e.g.,vaginal mucosal microbiota) of a female subject, the cervical microbiotaof a female subject, penile microbiota (e.g., penile skin microbiotasuch as the foreskin microbiota or the urethral meatus microbiota) of amale subject, microbiota of the genital tissue of an intersexindividual, microbiota of a non-binary gendered individual, or anycombination thereof. Recently, species overlap between rectal andurinary microbiome species in an individual have been observed. Forexample, the bacteria of the genital region have been found to representa continuum between organs of excretion and reproduction as discussed Y.Govender, et al., Front Cell Infect Microbiol 9 (2019): 133, herebyincorporated by reference in its entirety and particularly in relationto the urinary microbiomes disclosed therein. These bacteria of thegenital region are referred to herein as the anogenital and/orurogenital microbiomes.

As used herein, “genital probiotic bacteria” refer to live bacteria,which when administered in adequate amounts to the vagina or penisconfer a health benefit (e.g., such as those described herein) to thehost subject.

“Intersex” as used herein refers to an individual who has chromosomes,gonads, genitals, sex hormones, or other sex characteristics that can'tbe categorized as exclusively male or female.

As used herein, “non-binary gender” refers to an individual whose genderidentity isn't exclusively male or female.

As used herein, the term “vaginal microbiota” or “vaginal flora” refersto the collective microorganisms that normally colonize the vulva,clitoris, vestibule, and vagina and are non-pathogenic. In general, thebeneficial vaginal microbiota is primarily comprised of differentstrains of Lactobacillus (or related acid-producing bacterial types),which produce lactic acid to keep the vaginal ecosystem at a tightlycontrolled acidic environment (e.g., pH of about 3.5-5.5 or 3.5-5)during much of a woman's monthly cycle in reproductive aged women. Anexemplary vaginal microbiome includes a dominance of healthyLactobacillus species including: Lactobacillus jensenii, Lactobacillusgasseri, and Lactobacillus crispatus. Other beneficial species mayinclude Lactobacillus acidophilus, Lactobacillus plantarum,Lactobacillus fermentum, Lactobacillus brevis, Lactobacillus casei,Lactobacillus delbrueckii, Lactobacillus vaginalis, Lactobacillussalivarius, Lactobaccillus reuteri, and Lactobacillus rhamnosus.Additional bacterial species often present in low numbers in the vaginalregion of healthy women include: Lactobacillus iners, and species fromthe genuses Prevotella, Megasphaera, Colstridium, Baccilus, Gardnerella,Sneathia, and Mycoplasma. Additionally, acid producing bacteria that maybe part of the normal vaginal microbiota in some women includingLactobacillus iners, and species of Prevotella, Atopobium, Leptotrichia,Leuconostoc, Megasphaera, Pediococcus, Streptococcus, and Weissella. Thespecies found in normal vaginal microbiota can vary depending on age andethnicity. In some ethnicities and life stages, the acid-producing,healthy vaginal microbiome may not be primarily comprised ofLactobacillus spp. Evidence has shown that Lactobacillus spp. dominanceis associated with optimal reproductive and genital health outcomes inwomen of all ages. Various microbiota (and their connection with bothvaginal and non-vaginal health) are disclosed in J. Si, et al., CellHost & Microbe 21 (2017): 97-105, hereby incorporated by reference inits entirety, and particularly in relation to vaginal microbiotaincluding Lactobacillus and Prevotella species.

As used herein, the term “reproductive tract microbiota” refers to thecollective microorganisms that normally colonize the female upperreproductive tract, including the cervix, uterus, Fallopian tubes, andovaries, and are non-pathogenic. In certain embodiments, the femalereproductive tract microbiota can also be comprised of species from thegenus Pseudomonas, Acinetobacter, Vagococcus, Sphingobium,Erysipelothrix, Facklamia or Prevotella.

As used herein, the term “penile microbiota” or “penile flora” refers tothe collective microorganisms that normally colonize the penis,foreskin, and distal urethra which are non-pathogenic. The penisincludes the penile shaft and distal glans, which includes the glans,glans coronal, meatus urethralis, fossa navicularis, frenulum, coronalsulcus, and foreskin. In certain embodiments, the penile microbiotacomprises Lactobacillus crispatus, Lactobacillus jensenii, Lactobacillusiners, Lactobacillus gasseri, Streptococcus, non-pathogenic Prevotella,Corynebacteria, Staphylococcus, Anaerococcus, Peptoniphilus, Finegoldia,Porphyromonas, Propionibacterium, Delftia, Bifidobacterium, Clostridium,non-pathogenic Pseudomonas, or any combination thereof. In certainembodiments, the penile microbiota of a male subject reflects thevaginal microbiota and/or reproductive tract microbiota of a femalesubject, wherein the male subject and female subject are members of asexual dyad. The species found in normal penile microbiota can differbetween circumcised and uncircumcised subjects. For example, penilemicrobiota may also include sperm microbiota such as those disclosed inD Baud, et al., Frontiers in Microbiol 10 (2019): 234, herebyincorporated by reference in its entirety and particularly in relationto beneficial seminal microbiota including Lactobacillus species.

As used herein, the term “anogenital region” refers to the region of theanus and the genitalia. In certain embodiments, the female anogenitalregion comprises the cervix, vagina, vulva, clitoris, urethral meatus,urethral meatus, vulval vestibule, perineum, and/or anus. In certainembodiments, the male anogenital region comprises the penis, base of thepenis, foreskin, urethral meatus, scrotum, perineum, and anus. As usedherein, the term “urogenital region” refers to the region of the distalurinary tract and the genitalia. In certain embodiments, the femaleurogenital region comprises the cervix, vagina, vulva, clitoris,introitus, urethral meatus, urethral fold, vulval vestibule, and/orperineum. In some subjects, the anogenital and/or urogenital regions maybe indistinct, intersex, or transitioning from male to female or femaleto male due to iatrogenic (e.g., surgery or hormone therapy) ornatural/genetic causes.

As used herein, the term “genital tissues” refers to living cells foundin the anogenital and/or urogenital regions. Genital tissues include,but are not limited to epithelial surface cells (e.g., skin), mucosalcells, immune cells, nerve cells, blood cells, connective tissue cells,and neoplastic cells of the vulva, clitoris, vagina, vestibule, vulvalvestibule, urethral meatus, penis, foreskin, distal urethra, andscrotum. Since sperm cells exit the penis and are often deposited ongenital tissue (e.g., vagina), genital tissues also include semen andsperm cells.

As used herein, the term “genital fluid” refers to secretions from thebody that naturally occur in and around genital tissues. Genital fluidsinclude, but are not limited to cervical and vaginal secretions(together referred to as “cervico-vaginal fluids” (CVF)), semen, smegma,seminal fluid, urethral secretions, epithelial and mucosal coatings,menses flow, post-partum lochia, and other fluids naturally occurring inand around the vagina, vulva, clitoris, penis, foreskin and scrotum.

As used herein, the term “reproductive cycle” or “menstrual cycle”refers to hormone changes required for the production of an oocyte(ovarian cycle) and preparation of the uterus for pregnancy (uterinecycle). The reproductive cycle includes the time of fertilityimmediately before and after ovulation as well as the period whenconception is not possible due to the lack of a viable egg.

“Effective amount” or “therapeutically effective amount” refers to thatamount of a composition of this disclosure which, when administered to asubject, such as a human, is sufficient to effect a desired biologicaleffect or treatment, including any one or more of: (1) hydrating theurogenital and/or anogenital region; (2) lubricating the urogenitaland/or anogenital region; (3) cleaning the urogenital and/or anogenitalregion; (4) decreasing irritation or inflammation of the urogenitaland/or anogenital region; (5) maintaining, supporting, or enhancinghealthy genital and reproductive tract microbiota; (6) supporting orenhancing genital tissue function; (7) supporting or enhancing genitalfluid function; (8) supporting or enhancing sperm viability or function;(9) correcting or restoring optimal genital pH (e.g., to from about 4 toabout 5, to about 4.5); (10) decreasing bothersome genital symptoms suchas irritation; or any combination thereof. The amount of a compositionof this disclosure that constitutes an “effective amount” will varydepending on the formulation, the condition being treated and itsseverity, the manner of administration, the duration of treatment, orthe age or sex of the subject to be treated, but can be determinedroutinely by one of ordinary skill in the art having regard to his ownknowledge and to this disclosure.

As used herein, the term “subject” refers to any organism to which acomposition in accordance with the disclosure may be administered, e.g.,for experimental, diagnostic, prophylactic, and/or therapeutic purposes.Typical subjects include any animal (e.g., mammals such as mice, rats,rabbits, non-human primates, and humans, etc.). A subject in needthereof is typically a subject for whom it is desirable to treat adisease, disorder, or condition as described herein (and in particular,treatment of a disease, disorder or condition relating to dysbiosis ofthe urogenital and/or anogenital regions). For example, a subject inneed thereof may seek or be in need of treatment, require treatment, bereceiving treatment, may be receiving treatment in the future, or ahuman or animal that is under care by a trained professional for aparticular disease, disorder, or condition.

As used herein, the phrase “pharmaceutically acceptable” generally safefor ingestion or contact with biologic tissues at the levels employed.Pharmaceutically acceptable is used interchangeably with physiologicallycompatible. It will be understood that the pharmaceutical compositionsof the disclosure include nutraceutical compositions (e.g., dietarysupplements) unless otherwise specified.

The compounds described herein may be present as a pharmaceuticallyacceptable salt. Typically, salts are composed of a related number ofcations and anions (at least one of which is formed from the compoundsdescribed herein) coupled together (e.g., the pairs may be bondedionically) such that the salt is electrically neutral. Pharmaceuticallyacceptable salts may retain or have similar activity to the parentcompound (e.g., an ED₅₀ within 10%) and have a toxicity profile within arange that affords utility in pharmaceutical compositions. For example,pharmaceutically acceptable salts may be suitable for use in contactwith the tissues of humans and animals without undue toxicity,irritation, allergic response and are commensurate with a reasonablebenefit/risk ratio. Pharmaceutically acceptable salts are described in:Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977 and inPharmaceutical Salts: Properties, Selection, and Use, (Eds. P. H. Stahland C.G. Wermuth), Wiley-VCH, 2008. Salts may be prepared frompharmaceutically acceptable non-toxic acids and bases includinginorganic and organic acids and bases. Representative acid additionsalts include acetate, adipate, alginate, ascorbate, aspartate,benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate,camphorsulfonate, citrate, cyclopentanepropionate, dichloroacetate,digluconate, dodecyl sulfate, ethanesulfonate, formate, fumarate,glucoheptonate, glutamate, glycerophosphate, hemisulfate, heptonate,hexanoate, hippurate, hydrobromide, hydrochloride, hydroiodide,2-hydroxy-ethanesulfonate, i sethionate, lactobionate, lactate, laurate,lauryl sulfate, malate, maleate, malonate, mandelate, methanesulfonate,mucate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate,palmitate, pamoate, pantothenate, pectinate, persulfate,3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate,succinate, sulfate, tartrate, thiocyanate, toluenesulfonate,undecanoate, and valerate salts. Representative basic salts includealkali or alkaline earth metal salts include sodium, lithium, potassium,calcium, and magnesium, aluminum salts, as well as nontoxic ammonium,quaternary ammonium, and amine cations, including, but not limited toammonium, tetramethylammonium, tetraethylammonium, methylamine,dimethylamine, trimethylamine, triethylamine, caffeine, and ethylamine.

Pharmaceutically acceptable acid addition salts of the disclosure can beformed by the reaction of a compound of the disclosure with an equimolaror excess amount of acid. Alternatively, hemi-salts can be formed by thereaction of a compound of the disclosure with the desired acid in a 2:1ratio, compound to acid. The reactants are generally combined in amutual solvent such as diethyl ether, tetrahydrofuran, methanol,ethanol, iso-propanol, benzene, or the like. The salts normallyprecipitate out of solution within, e.g., one hour to ten days and canbe isolated by filtration or other conventional methods.

Prodrugs are typically compounds that may be converted underphysiological conditions or by solvolysis to a biologically activecompound of the invention. Prodrug may refer to a metabolic precursor ofa compound of the invention that is pharmaceutically acceptable. Aprodrug may be inactive when administered to a subject in need thereof,but is converted in vivo to an active compound of the invention.Prodrugs are typically rapidly transformed in vivo to yield theindicated compound, for example, by hydrolysis in blood. The prodrugcompound often offers advantages of solubility, tissue compatibility ordelayed release in a mammalian organism as described in Bundgard, H.,Design of Prodrugs (1985): 7-9, 21-24 (Elsevier, Amsterdam), Higuchi,T., et al., ACS Symposium Series, Vol. 14, and Bioreversible Carriers inDrug Design, Ed. Edward B. Roche, American Pharmaceutical Associationand Pergamon Press, 1987, each of which are hereby incorporated byreference in their entirety.

Compositions

The present disclosure provides topical, isotonic compositionscomprising a prebiotic oligosaccharide, a metal co-factor, and bornylacetate (e.g., the composition comprises an essential oil comprisingbornyl acetate), at a pH ranging from about 3.5 to about 8. Suchcompositions have no or minimal irritation to the epithelial tissues ofthe urogenital and/or anogenital regions; no or minimal disruption ofnatural mucus secretions and mucins of the urogenital and/or anogenitalregions; minimal or no inhibition on the normal, non-pathogenicmicrobiota of the urogenital and/or anogenital regions; do not promotegrowth of pathogenic bacteria of the urogenital and/or anogenitalregions; limit transfer of pathogenic bacteria from a first member of adyad to a second member of a dyad; enhance transfer of beneficialmicrobiota species from a first member of a dyad to a second member of adyad; or any combination thereof.

Distinct genital microbiome states and metabolic profiles have beenidentified that correlate with health and conversely with genitaldisease (e.g., penile or vaginal disease), fertility, poor birthingoutcomes, HIV susceptibility, HPV susceptibility, sexually transmittedinfections, autoimmune disease, and cancer risk. Systemic risk diseasessuch as cognitive impairment/dementia may also be associated with thechronic inflammation and dysbiosis of the genital microbiome as shown inR. Muzambi, et al., BMJ Open 9 (2019): e030874, hereby incorporated byreference in its entirety, and particularly in relation to theconnection between bacterial infections and dementia. Additionally,diseases such as Type 1 diabetes in offspring have also been associatedwith the genital microbiome of the mother as disclosed in M. Tejesvi, etal., Sci Rep 9 (2019): 959, hereby incorporated by reference in itsentirety and particularly in relation to the connection between thevaginal microbiome and Type 1 diabetes in members of a dyad.

Therefore, it is important to consider the genital microbiome and itsimplications on human health in the development of genital and sexualhealth products and therapeutics. In general, the healthy human vaginalmicrobiome, which is dominated by predominantly Lactobacilli species,maintains and utilizes a carbohydrate-based, antioxidant-richenvironment (Chen et al. Nat Commun 8:875, 2017, hereby incorporated byreference in its entirety). These Lactobacilli produce distinctmetabolites that stimulate epithelial mucus production, regulate immunecell reactivity, control pathogenic biofilm and biogenic amineformation, and prevent the colonization of the vaginal tract bypathogenic bacteria. The penile microbiome may reflect, and iscorrelated to, the vaginal microbiome of a female sexual partner.Dysbiosis, which refers to impaired or imbalanced microbiota, can leadto outgrowth of pathogenic, or dysbiotic, microorganisms. Dysbioticorganisms tend to prefer amino acids or proteins as energy substratesrather than carbohydrates, produce an antioxidant-poor environment(which is high in cytokine active metabolites) degradation products suchas biogenic amines, and certain anti-oxidant reducing agents. Theseproducts often lead to a variety of bothersome symptoms and pathologiesfrom odor to itching, inflammation, biofilm production (which mayprotect pathogens from the host immune system), mucin secretiondegradation, and higher risks of viral infections and cancer. Certainlife events cause an individual to be more susceptible to dysbiosis,including: ovulation, menses, birth, puberty, pregnancy, sexualinitiation, non-circumcision of the male, genital mutilation the female,diseases (e.g., diabetes), introduction of new sexual partners,post-partum, involution of the uterus, times of immunosuppression,menopause or andropause, and cancer therapy. The present disclosure isrelated to compositions and methods which do not elicit or minimize theoccurrence of such responses and/or maintain the healthy microbiome ofthe anogenital and/or urogenital microbiomes.

The pathogenesis of vaginal and penile chronic dysbiosis, rely on theproduction of bioactive amines. A few species of dysbiotic bacteria(e.g., the bacteria present in a microbiome under dysbiosis) produce themost significant amounts of biogenic amines such as agmatine,putrescine, cadaverine, and tyramine, which increase vaginal pH and makethe vaginal microbiome more inviting to other dysbiotic species. Thesespecies may also produce bacterial endotoxins such aslipopolysaccharides which have been linked to diseases, disorders, andconditions such as various cancers and dementia. Vaginal dysbiosis isoften linked to concurrent male partner penile dysbiosis. This shift inmetabolic states from a glycogen/carbohydrate to amino acid-richenvironment results in a shift in vaginal pH away from the beneficialacidic state (pH about 4.5) to a more alkaline state. Such shift isinitiated and perpetuated by specific bacteria including the genera:pathogenic Prevotella, Eggerthellia, Gardnerella, Atopobium,Megasphaera, Mobiluncus, Mageeibacillus, Gemella, Veillonella, Snethia,Mycoplasma, and some Clostridium species. It is important to note thatsome of these bacteria are not most abundant or solely present in adysbiotic microbiome, confirming that the metabolic activity and theecological role that a microbe plays in an environment is more importantthan its abundance. These dysbiotic bacteria produce metabolites thatresult in increased genital pH, offensive odors, biofilm production thatinterferes with host immune function, increased epithelial inflammation(which may result in symptoms such as burning, itching, and/or pain),increased mucin degradation (which may result in symptoms such as pain,roughness of the skin, and/or irritation), increased oxidative stress,cytokine and inflammasome production, and decreased growth capacity ofhealthy microbiome organisms. These conditions leave the individualsusceptible to the development of further dysbiosis and disease states,including increased risk of poor reproductive outcomes (e.g., failedfertilization, failed embryo implantation, poor fetal growth, deliverycomplications), pain during sex, sexually transmitted diseases (such ashigh risk Human Papilloma virus (HPV) infection and persistence, genitalHerpes Simplex Virus (HSV) infection, and Human Immunodeficiency Virus(HIV) infection), auto-immune conditions, cancer, and systemic disease(e.g., cognitive impairment). These conditions also have a social costof embarrassment and social withdrawal for women with odor and dischargeand for men with odor and visual roughness or inflammation of penileskin. These social aspects of genital dysbiosis are rarely discussed andunder treated.

The genital microbiome of individuals may also impact sexual partners oroffspring. Couples with a female partner having bacterial vaginosis (BV)show high levels of shared pathogenic bacteria with the male partner(over 17 species shared). In contrast, healthy couples also sharebacterial species, but have a more narrow range (e.g., less diversity)of microorganisms (e.g., commonly 4 strains of bacteria), includingLactobacillus. A menopausal woman may experience chronic bacterialdysbiosis, which favors the growth of pathogenic bacteria. Thisdysbiosis may be transmitted to the sexual partner of the menopausalwomen. For male sexual partners, penile microbiota has an important rolein prostate health and disease. Penile dysbiosis of the male sexualpartner may induce chronic inflammation, which may predispose the malesexual partner to subfertility, prostatitis, chronic pelvic painsyndrome, benign hyperplasia, as well as prostate cancer (Porter et al.,Prostate Cancer Prostatic Dis. 21:345, 2018). The penile microbiome alsoaffects systemic hormone levels (Id.). In another example oftransmission of dysbiosis, an uncircumcised man with chronic peniledysbiosis can transmit dysbiosis to a female sexual partner, resultingin recurring bacterial vaginosis. Currently, women with bacterialvaginosis may undergo repeated rounds of antimicrobial therapy. However,without concurrent treatment of the male sexual partner, relapse andcontinued retreatment of the woman have become routine.

Natural vaginal cleaning and protection of the woman from pathogensencountered during vaginal intercourse depends on several factorsincluding: 1) an acidic environment (e.g., pH from about 4.0 to about4.5) that facilitates constant lysis and shedding of the outer layer ofvaginal epithelial cells; 2) cervico-vaginal fluids (CVF) forming in thevaginal canal that have a low salt/ion concentration compared to bloodand fluids bathing inner tissues, which further facilitates lysis andshedding of the outer most vaginal epithelial cell lining, and 3) theacidic mucin-coated lining of the vaginal canal formed by mucinparticles embedded in the vaginal lining, which shed to the externalcanal. Constant turn-over of the vaginal lining results in out-flow fromthe vagina, facilitating removal of pathogens before they can establishin the woman's body. However, the normally low pH and low saltenvironment of the vaginal canal environment can be toxic to sperm. Thistoxicity is mitigated during ovulation by a shift in the cervical canalderived secretions to a more neutral pH that supports sperm viabilityand motility (e.g., pH from about 6 to about 7) and by the deposition ofsperm in semen during ejaculation. Semen has a more alkaline pH (e.g.,pH of about 8) and a higher salt concentration (3 times that ofcervico-vaginal fluids (CVF) and 1.5 times that of blood). The admixingof semen and cervical fluids during vaginal intercourse at ovulationprovides a neutral pH and a balanced salt environment in the vaginalvault. This environments aids in protecting sperm for their transitthrough the female reproductive tract (vagina and cervix). However,vaginal dysbiosis can occur if the acidic, low salt environment of thevaginal canal is not rapidly restored following the fertile phase of thewoman's cycle.

The impact of the shared microbiome of a sexual dyad is particularlynotable during reproduction. Sexual dyads that are trying to conceiveoften have disrupted genital microbiota. Reproduction in humans requiresan elegant interaction between the woman's host defenses in the vaginaand the ejaculate of the man, which provides sperm cell transport andsurvival, to occur, so that fertilization of the egg follows. Subfertilewomen have increased levels of vaginal dysbiosis and poor vaginalmicrobiome resilience. Penile dysbiosis can interfere with sperm and/orsemen quality. It is critical that a healthy genital microbiome berestored and maintained in couples with poor fertility. Furthermore,bacterial vaginosis and other forms of dysbiosis can increase risk ofsexually transmitted diseases; cancers (e.g., cervical cancer); pelvicinflammatory disease; infertility; miscarriage; ectopic pregnancy; andpoor reproductive and birth outcomes. Bacterial vaginosis is alsoassociated with upper reproductive tract dysbiosis, which can affect theFallopian tubes and uterus. Upper reproductive tract dysbiosis canresult in infertility; miscarriage ectopic pregnancy; and pregnancycomplications, including premature rupture of membranes, preterm labor,premature birth, low birth weight, chorioamnionitis, and endometritis.Pregnant women with active bacterial vaginosis have a higher risk ofCesarean-section.

Additionally, the microbiome of the fetus and newborn is shaped by themother-fetus, mother-child dyad. The initial fetal microbiome isdeveloped from the maternal reproductive tract microorganisms. Thenewborns' microbiome is seeded at birth upon exposure to maternalvaginal, fecal, and skin microbiota. Disrupting the mother-to-newborntransmission of microbiome by C-section delivery may increase the riskof celiac disease, asthma, Type 1 diabetes, and obesity in the offspring(see, Mueller et al., Trends Mol. Med. 21:109, 2015).

Genital dysbiosis is also common in individuals undergoing genderreassignment surgery. Active and ongoing management of newly createdgenitalia requires daily cleaning and intervention to assist in healthygenital biome development of these newly created regions (see, Weyers etal., BMC Microbiol. 9:102, 2009).

Additionally, urinary incontinence, diabetes, genital lichen sclerosus,interstitial cystitis, and autoimmune conditions can increase genitaldysbiosis (Thomson, J Reprod. Med. 50:513, 2005; Alam et al. Immune Netw14:7, 2014). Individuals with such diseases can be very sensitive todisease exacerbation via ingredients or chemicals in formulationscurrently used for routine genital care and intimacy as disclosed in AChung, et al., World J Urol (2019): 1-5.

Overall, these diseases, disorders, and/or conditions are poorlycontrolled and poorly managed in many populations. Furthermore, theetiology of certain systemic disease conditions such as dementia (e.g.,associated with incontinence and diabetes), and cancer (e.g., associatedwith squamous cell cancer in lichen patients) are connected withdysbiosis of the anogenital and/or urogenital regions.

The present disclosure provides genital products that can optimizegenital health and modulate the metabolism of the genital microbiome.The topical, isotonic compositions of the present disclosure includecomponents for supporting growth, dominance, resilience, and metabolismof beneficial lactic-acid producing bacteria and components forinhibiting dysbiotic bacterial growth and metabolism. By influencing themetabolism of the genital microbiome, the growth and metabolic activityof beneficial bacterial species (e.g., Lactobacillus species) can beenhanced, and the growth and metabolism of bacterial species that leadto genital dysbiosis and disease can be inhibited to restore andmaintain genital health. By treating all the members of dyads, triads,and higher order social groups of significance, the health of all themembers of dyads, triads, or larger social groups can also be improved.Furthermore, people in high stress, crowded, or unhygienic situations,such members of the military, incarcerated individuals, people living indormitories or shelters, people in man-made or natural disasters canalso form a dyad or other social grouping with extensive sharing ofanogenital microbiota through daily interactions. People living in suchsettings can benefit from the improved cleaning and personal careoffered by the compositions of the present disclosure.

Topical, isotonic compositions of the present disclosure comprise aprebiotic oligosaccharide in an amount ranging from about 0.001% toabout 5% by weight of the composition. Prebiotic oligosaccharide refersto oligosaccharides substrates that induce the growth or activity ofbeneficial microorganisms of the microbiota, e.g., genital microbiota.In certain embodiments, a prebiotic is non-digestible and resistant tobreakdown by stomach acid and enzymes in the human gastrointestinaltract, selectively fermented by genital microbiota (e.g., beneficialgenital microbiota), selectively target and stimulate the growth andactivity of specific genital microbiota (e.g., healthy and/or beneficialgenital microbiota), or any combination thereof.

One or more prebiotics may be added to composition. Suitable prebioticsmay include oligosaccharides, particularly galactooligosaccharide (GOS),palatinoseoligosaccharide, soybean oligosaccharide,gentiooligosaccharide, xylooligomers, non-degradable starch,lactosaccharose, lactulose, lactitol, maltitol, polydextrose, or thelike. Examples of prebiotic oligosaccharides that may be used in thecompositions of the present disclosure include raffinose, lactulose,trehalose, galactooligosaccharide, alpha-glucan oligosaccharide,beta-glucan oligosaccharide, maltose, xylose, fructooligosaccharide,isomaltooligosaccharide, inulin, pectin, or any combination thereof. Incertain embodiments, a prebiotic oligosaccharide does not includexylose.

Topical, isotonic compositions of the present disclosure may alsocomprise a metal co-factor. Metal co-factors may be metallic ions, orsalts thereof, which often act as a catalyst for an enzyme's activity.Specifically, the metal co-factors may assist enzymes involved inglycosylation of proteoglycans, such as glycosaminoglycans, which areinvolved in a variety of physiological functions including barrierimmune protection, epithelial hydration and providing viscosity tonatural bodily fluids, such as cervico-vaginal secretions, penileforeskin secretions, smegma, distal urethral sections, and semen. Themetal co-factor may be requisite in some conditions for Lactobacillusgrowth. Pathogenic bacteria may sequester such metal co-factors, as anact of establishing dominance over beneficial genital microbiota. Themetal co-factor may be present in an amount ranging from about 0.0001%to about 0.1% by weight of the composition. A metal co-factor maycomprise zinc, selenium, manganese, cobalt, iron, copper, includingsalts thereof, or any combination thereof. The metal co-factor may beadded to the composition as a salt of the co-factor metallic ioncomprising a counterion, for example, the chloride salt. For example,the metal co-factor may comprise Mn²⁺, Mn³⁺, Mn⁴⁺, Mn⁶⁺, Mn⁷⁺, Zn¹⁺,Zn²⁺, Se²⁺, Se⁴⁺, Se⁶⁺, Co²⁺, Co³⁺, Fe²⁺, Fe³⁺, Cu¹⁺, Cu²⁺, orpharmaceutically acceptable salts (e.g. the chloride salt) thereof, orhydrates of any of the foregoing. In some embodiments, the metalco-factor is manganese (II) chloride. It will be understood that somehydrates of these metallic salts may dissociate to form themetal-cofactor in the topical isotonic solutions.

Bornyl acetate is a compound commonly found in pine needles, valerianroot, fir needles, hemlock, cypress, rosemary, and occasionally juniperberries and spearmint. Bornyl acetate has acetylcholinesterase enzymeinhibitory, anti-inflammatory, and analgesic activity as shown forseveral indications in described in L Yang, et al., Biomed Pharmacother103 (2018): 234-239, S. Lu et al. Biomed Res Int 2018 (2018): 3589874, TZhang et al. Front Pharmacol 8 (2017): 786, and D Szwajgier et al., CurrAlzheimer Res 16 (2019): 963, each hereby incorporated by reference intheir entirety. In some embodiments, the pharmaceutical compositioncomprises bornyl acetate or pharmaceutically acceptable salts orprodrugs thereof. In some embodiments, the pharmaceutical compositioncomprises borneol or pharmaceutically acceptable salts thereof orprodrugs (e.g., ester prodrugs) of any of the foregoing. Thecompositions of the present disclosure may comprise more than about0.0001% bornyl acetate by weight of the composition (e.g., more thanabout 0.00025% bornyl acetate, from about 0.0001% to about 1% bornylacetate, from about 0.0001% to about 0.5% bornyl acetate, from about0.0001% to about 0.3% bornyl acetate, from about 0.0001% to about 0.2%bornyl acetate, from about 0.0001% to about 0.1% bornyl acetate, fromabout 0.0001% to about 0.001% bornyl acetate, from about 0.001% to about0.01% bornyl acetate, from about 0.01% to about 0.1% bornyl acetate,from about 0.1% to about 1% bornyl acetate, from about 0.01% to about0.3% bornyl acetate, from about 0.1% to about 0.3% bornyl acetate, fromabout 0.00025% to about 0.20% bornyl acetate, 0.0005% to about 0.20%bornyl acetate, from about 0.0001% to about 0.15% bornyl acetate, fromabout 0.00025% to about 0.15% bornyl acetate, from about 0.001% to about0.06% bornyl acetate) by weight of the composition. In some embodiments,the composition may comprise less than about 0.3% (e.g., less than about0.15%, less than about 0.015%, from about 0.0001% to about 0.015%, fromabout 0.00025% to about 0.015%) bornyl acetate by weight of thecomposition. In some embodiments, the compositions of the presentdisclosure may comprise more than about 0.0001% borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing (e.g., bornyl acetate) by weight of the composition (e.g.,more than about 0.00025% borneol, pharmaceutically acceptable saltsthereof, or prodrugs of any of the foregoing, from about 0.0001% toabout 1% borneol, pharmaceutically acceptable salts thereof, or prodrugsof any of the foregoing, from about 0.0001% to about 0.5% borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing, from about 0.0001% to about 0.3% borneol, pharmaceuticallyacceptable salts thereof, or prodrugs of any of the foregoing, fromabout 0.0001% to about 0.2% borneol, pharmaceutically acceptable saltsthereof, or prodrugs of any of the foregoing, from about 0.0001% toabout 0.1% borneol, pharmaceutically acceptable salts thereof, orprodrugs of any of the foregoing, from about 0.0001% to about 0.001%borneol, pharmaceutically acceptable salts thereof, or prodrugs of anyof the foregoing, from about 0.001% to about 0.01% borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing, from about 0.01% to about 0.1% borneol, pharmaceuticallyacceptable salts thereof, or prodrugs of any of the foregoing, fromabout 0.1% to about 1% borneol, pharmaceutically acceptable saltsthereof, or prodrugs of any of the foregoing, from about 0.01% to about0.3% borneol, pharmaceutically acceptable salts thereof, or prodrugs ofany of the foregoing, from about 0.1% to about 0.3% borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing, from about 0.00025% to about 0.20% borneol, pharmaceuticallyacceptable salts thereof, or prodrugs of any of the foregoing, 0.0005%to about 0.20% borneol, pharmaceutically acceptable salts thereof, orprodrugs of any of the foregoing, from about 0.0001% to about 0.15%borneol, pharmaceutically acceptable salts thereof, or prodrugs of anyof the foregoing, from about 0.00025% to about 0.15% borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing, from about 0.001% to about 0.06% borneol, pharmaceuticallyacceptable salts thereof, or prodrugs of any of the foregoing) by weightof the composition. In some embodiments, the composition may compriseless than about 0.3% borneol, pharmaceutically acceptable salts thereof,or prodrugs of any of the foregoing (e.g., less than 0.15% borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing, less than 0.015% borneol, pharmaceutically acceptable saltsthereof, or prodrugs of any of the foregoing, from about 0.0001%borneol, pharmaceutically acceptable salts thereof, or prodrugs of anyof the foregoing to about 0.015% borneol, pharmaceutically acceptablesalts thereof, or prodrugs of any of the foregoing, from about 0.00025%borneol, pharmaceutically acceptable salts thereof, or prodrugs of anyof the foregoing to about 0.015% borneol, pharmaceutically acceptablesalts thereof, or prodrugs of any of the foregoing), by weight of thecomposition.

Borneol, bornyl acetate, pharmaceutically acceptable salts thereof, orprodrugs of any of the foregoing typically have at least one chiralcenter. The compositions may comprise the enatiomerically pure compound,enantiomeric mixtures of an indicated compound, or racemic mixtures ofthe enantiomer. For example, bornyl acetate may be present as (+)-bornylacetate, (−)-bornyl acetate, racemic mixtures thereof, or enantiomericmixtures thereof (e.g., a weight ratio of (+)-bornyl acetate to(−)-bornyl acetate of from about 100:1 to about 1:100 or about 50:1 toabout 1:50 or about 25:1 to about 1:25 or about 10:1 to about 1:10).

In some embodiments, topical, isotonic compositions of the presentdisclosure may also comprise an essential oil comprising bornyl acetate,a monoterpene ester, in an amount ranging from about 0.005% to about0.5% by weight of the composition. In certain embodiments, the essentialoil comprising bornyl acetate comprises essential oil from Juniperuscommunis, Juniperus occidentalis, Juniperus scopulorum, Abies sibirica,Abies alba, Abies balsamea, Abies fraseri, Abies grandis, Abiesspectabilis, Abies koreana, Abies procera, Abies nordmanniana, Abiesmagnifica, Abies pinsapo, Abies lasiocarpa, Abies concolor, Pseudotsugamenziesii, Ambrosia trifida, Pinus mugo, Romanian solidago, Ribesnigrum, Laurus nobilis, Rosmarinus officinalis, or any combinationthereof. In some embodiments, the essential oil comprising bornylacetate comprises bornyl acetate in an amount of at least about 5% byweight of the essential oil. In some embodiments, the essential oilcomprising bornyl acetate comprises bornyl acetate in an amount rangingfrom about 5% to about 30% or from about 10% to about 30% by weight ofthe essential oil. Accordingly, in some embodiments, the topical,isotonic compositions may comprise more than about 0.00025% bornylacetate (e.g., from about 0.00025% to about 0.15% bornyl acetate,0.0005% to about 0.15% bornyl acetate) by weight of the composition. Insome embodiments, the topical isotonic compositions may comprise from0.0001% bornyl acetate to about 0.3% bornyl acetate by weight of thecomposition. In certain implementations, the topical isotoniccompositions may comprise an essential oil comprising bornyl acetatesuch that from 0.0001% bornyl acetate to about 0.3% bornyl acetate(derived from the essential oil) by weight of the composition.

In certain embodiments, topical, isotonic compositions of the presentdisclosure further comprise a bisabolene. The bisabolene may be in anamount ranging from about 0.0001% to about 0.1%. Bisabolene hasanti-inflammatory, wound healing, skin strengthening, anti-tumor and/oranalgesic activity. In certain embodiments, bisabolene is obtained fromCommiphora guidotti, Pallines spinosa, Platanus chiapensis, Platanusgentryi, Platanus kerrii, Platanus mexicana, Platanus oaxacana, Platanusoccidentalis, Platanus orientalis, Platanus racemosa, Platanusrzedowskii, Platanus wrightii, Platanus acerifolia, or any combinationthereof.

In certain embodiments, topical, isotonic compositions of the presentdisclosure further comprise a flavonoid. The flavonoid may be in anamount ranging from about 0.001% to about 0.1%. Flavonoids possess awide range of biological and pharmaceutical activities, includingantioxidant and anti-inflammatory activities. In certain embodiments,the flavonoid comprises catechin, epicatechin, rutin, luteolin,apigenin, kaempherol, myricetin, quercetin, quercitrin, naringin,naringenin, hesperetin, hesperidin, taxifolin, genistin, genistein,daidzein, cyanidin, apigenidin, tangeritin, fisetin, galangin,isorhamnetin, pachypodol, rhamnazin, pyranoflavonol, fluranoflavonol,eriodictyol, homoeriodictyol, taxifolin, gallocatechin, catechin3-gallate, gallocatechin 3-gallate, epigallocatechin, epicatechin3-gallate, epigallocatechin 3-gallate, theaflavin, thearubigin,proanthocyanidin, dephinidin, malvidin, pelargonidin, peonidin,petunidin, isoflavone, glycitein, isoflavane, isoflavandiol, isoflavene,coumestan, pterocarpan, myricitrin, phloridzin, or any combinationthereof. In certain embodiments, the flavonoid comprises a citrusessential oil (e.g., Citrus reticulata) as disclosed in Y Yang, et al.,J Food Sc 12 (217): 2840-2846, hereby incorporated by reference in itsentirety, wherein monoterpene hydrocarbons comprise at least 75% or atleast 85% of the total composition of the essential oil. Monoterpeneconcentration may be measured using the method as described in Njorogeet al. (Journal of Essential Oil Research 18:659, 2006) herebyincorporated by reference in its entirety.

In certain embodiments, topical, isotonic compositions of the presentdisclosure may further comprise a sesquiterpene alcohol and/ormonocyclic sesquiterpene. The sesquiterpenes may be in an amount rangingfrom about 0.001% to about 0.1% by weight of the composition.Sesquiterpenes possess a wide range of biological and pharmaceuticalactivities, including increasing skin hydration, antimicrobial activity,and antifungal activity. In certain embodiments, the sesquiterpenes mayinclude nerolidol, an essential oil from Citrus aurantum var sp,bisabolol, patchoulol, alpha-santalol, zingiberene, or combinationsthereof. In some embodiments, the sesquiterpene alcohol or monocycliccomprises nerolidol, an essential oil from Citrus aurantum (e.g., Citrusaurantium var. amata) and/or Citrus bigaradia, bisabolol, patchoulol,alpha-santalol, zingiberene, or combinations thereof. In certainimplementations the sesquiterpene alcohol and/or monocyclicsesquiterpene is neroli oil.

In certain embodiments, topical, isotonic compositions of the presentdisclosure further comprise a biofilm inhibiting agent. The biofilminhibiting agent may be in an amount ranging from about 0.001% to about0.16%. In certain embodiments, the biofilm inhibiting agent comprisesLamiaceae essential oil, Garcinia extract, Eurycoma longifolia extract,or any combination thereof. Examples of Lamiaceae essential oil includeessential oil from Mentha spicata, Mentha pulegium, Mentha piperita,Mentha aquatica, Mentha arvensis, Mentha asiatica, Mentha australis,Mentha canadensis, Mentha cervina, Mentha citrata, Mentha crispata,Mentha dahurica, Mentha diemenica, Mentha laxiflora, Mentha, longifolia,Mentha requienii, Mentha sachalinensis, Mentha satureioides, Menthasuavenolens, Mentha vagans, Melissa officinalis, Monarda Fistulosa, orany combination thereof. Garcinia plants are sources of antioxidantxanthones, which possess anti-microbial and anti-inflammatoryproperties. Examples of xanthones include y-mangostin, garcinone-D,gartanin, and smeathxanthone. Examples of Garcinia extract includeextract from Garcinia mangostana, Garcinia travancorica, Garciniacambogia, Garcinia kola, Garcinia zeylanica, Garcinia xanthochymus, orany combination thereof.

In certain embodiments, topical, isotonic compositions of the presentdisclosure further comprise a cell membrane active phytoesterol. Thephytoesterol may be in an amount ranging from about 0.001% to about0.1%. Phytoesterols may inhibit the growth of pathogenic bacteria andhave the potential to inhibit the activity of pore-forming toxins suchas vaginolysin, leukotoxin, and other cholesterol dependent cytotoxinsfound in pathogenic strains of bacteria such as some Staphylococcus,Clostridium, and Gardnerella spp. In certain embodiments, thephytoesterols comprise apigenin, β-sitosterol, campesterol,brassicasterol, stigmasterol, sitosterol, or any combination thereof.Examples of phytoesterol sources include ginseng (Panax quinquefolium)seed extract, carrot, yam or coriander extract, ginger root extract,Mirabilis Jalap, or any combination thereof.

In certain embodiments, topical, isotonic compositions of the presentdisclosure further comprise a prebiotic spice extract. The prebioticspice extract may be in an amount ranging from about 0.001% to about0.02%. Exemplary prebiotic spice extracts include extract from ginger,black pepper, cayenne pepper, cinnamon, oregano, rosemary, turmeric, orany combination thereof.

In certain embodiments, topical, isotonic compositions of the presentdisclosure further comprise a viscosity-increasing agent. Viscosity is aproperty of liquids that is closely related to the resistance to flow.It may be defined by Couette flow, which is the laminar flow of aviscous fluid in the space between two parallel plates, one of which ismoving relative to the other. The flow is driven by virtue of viscousdrag force acting on the fluid and the applied pressure gradientparallel to the plates.

The topical, isotonic compositions may comprise one or more rheologyagents. In some embodiments, the composition may comprise one or morenon-cellulose based rheology agents. In some embodiments, the topical,isotonic composition comprises a rheology agent selected frompoloxamers, polybutene, polycarbophil, polyvinylalcohol,polyvinylpyrrolidone polymers, polyoxazoline polymers, and combinationsthereof.

In some embodiments, the compositions of the present disclosure furthercomprise one or more humectants such as glycerin, hexylene glycol,arabinogalactan, caprylyl glycol, or combinations thereof. In certainembodiments, the compositions may comprise a humectant that is anextract of a plant from the genus Monotropa (e.g., Monotropa hypopitys)such as those described in JP App No 2009191075 A to Yamada, herebyincorporated by reference in its entirety.

In some embodiments, the compositions of the present disclosure furthercomprise one or more anti-inflammatory agents and/or one or moresoothing agents. Specific anti-inflammatory and/or soothing agentsinclude ICAM inhibitors (e.g., CD11a, ezrin (EZR), CD18, glycyrrhetinicacid, pyrrolidinedithiocarbamate), NFκB inhibitors (e.g., (heterocyclicthiazole, lipoic acid, efalizumab,4-[(4-Methylphenyl)thio]thieno[2,3-c]pyridine-2-carboxamide, silibinin,stilbenes, (+)-epigalloylcatechin-gallate [(+)-EGCG]), cytokineinhibitors TSLP inhibitors, IL-25 inhibitors, IL-33 inhibitors, IL-1inhibitors, TNF inhibitors (e.g., TNF-α inhibitors, TNF-β inhibitors),quercetin and isoforms thereof (e.g., isoquercetin, etc.), non-steroidalanti-inflammatory drugs (e.g., aspirin), and extracts from plants of thegenus Vigna (e.g., Vigna Caracalla), extracts of plants from the genusRhododendron (e.g., Rhododendron aceae), extracts of plants from thesubfamily Monotropacease such as Allotropa virgate as disclosed in JP2009191075 to T Yamada, hereby incorporated by reference in itsentirety, and combinations thereof. In various embodiments, theanti-inflammatory agent and/or soothing may be present in an amount offrom about 0.0001 to about 10% (e.g. from about 0.001 to about 5%) byweight of the composition

The extracts may be prepared by enzymatic extraction, solventextraction, steam distillation, or any other method known in the art. Insome embodiments, at least one of the topical compositions of theinvention comprises an extract, obtained by steam distillation, of anyof the forgoing plants and biological materials (each one beingconsidered a distinct embodiment). In some embodiments, at least one ofthe topical compositions of the invention comprises an extract, obtainedby extraction with water (e.g., basic, neutral, or acid), of any of theforgoing plants and biological materials (each one being considered adistinct embodiment). The water of extraction may further include aco-solvent miscible with water, including lower alcohols (e.g., C₁₋₆),such as methanol, ethanol, isopropanol, propanol, butanol (typically,ethanol). In some embodiments, at least one of the topical compositionsof the invention comprises an extract, obtained by extraction with asolvent system comprising from about 5-95% (v/v) or 10-90% (v/v) or20-80% (v/v) or 40-60% (v/v) water (e.g., basic, neutral, or acid) andabout 5-95% (v/v) or 10-90% (v/v) or 20-80% (v/v) or 40-60% (v/v)ethanol, of any of the forgoing plants and biological materials (eachone being considered a distinct embodiment). In some embodiments, atleast one of the topical compositions of the invention comprises anextract, obtained by extraction with an organic solvent (e.g.,non-polar, polar aprotic, or polar protic), of any of the forgoingplants and biological materials (each one being considered a distinctembodiment). Suitable solvents include hexane and other C₁₋₁₂ or C₅₋₈hydrocarbons, lower alcohols, C₂₋₁₆ ethers (e.g., diethyl ether), C₃₋₁₂esters (e.g., ethyl acetate), C₂₋₁₂ (e.g., acetone, butanone), carbondioxide (liquid or supercritical) The biological extracts may be driedunder vacuum or atmospheric pressure to remove water and solvents ofextraction. The biological extracts may be dried by lyophilization. Thebiological extracts may be passed over activated carbon or charcoaland/or passed through filters and/or microfilters to remove bacteria andother biological materials.

In certain embodiments, compositions of the instant disclosure areformulated to have viscosity best suited for the target tissue (e.g.,anogenital region) and to mimic the properties of normal genital fluids.For example, compositions formulated as gels applied to mucous membranesmay be designed to have viscosity values consistent with or similar tonormal mucus, and exhibiting non-Newtonian, shear-thinning(pseudoplastic) flow properties. Standardized methodology forquantitative comparisons of over-the-counter vaginal products basedfeatures such as, stickiness, ropiness, peaking, rubberiness, thickness,smoothness, and slipperiness, are known in the art (Mahan et al.,Contraception, 84:184, 2011). In some embodiments, compositionsformulated as gels applied to mucous membranes may strengthen mucusquality and/or mucin coverage of the body surface. In certainimplementations, the compositions may stimulate the production orproffer an acidic barrier in the urogentical and/or anogenital regionwith improved muco-adhesion, which may increase the bioavailablity ofone or more active components of the composition and result in abeneficial impact on the genital microbiome as disclosed in N Peppas, etal., J Biomater Sci Polym Ed. 20 (2209): 1-20, hereby incorporated byreference in its entirety and particularly in relation to muco-adhesivecarrier development.

For compositions applied to skin (such as the vulva, perineum, or penis)or inside the vagina, a viscosity-increasing agent can be added in anamount that allows the composition to spread easily to form a thin layerwhen minimal physical pressure is applied, and to have adequateviscosity and shear-thinning properties so that the composition does not“run” off or out of the genital tissue upon topical application.Mucoadhesive formulations that are retained at the genital surface(e.g., vulvar, vaginal, penile, foreskin surface) for prolongedbiological activity are known in the art (reviewed by Khutoryanskiy,Macromol. Biosci. 11:748, 2011; Brooks, Front. Chem. 3:65, 2015).Muco-adhesive formulas must have polymer compositions that activelyadmix and interact with physiologic mucin and mucus of secretions. Somecommon gelling agents do not intertwine with natural mucins and aretherefore not muco-adhesive and are rapidly lost from the epithelium.

Compositions of the present disclosure may comprise aviscosity-increasing agent in an amount ranging from, for example, about0.05% to about 10% by weight of the composition. In certain embodiments,the viscosity enhancing agent comprises a tensioactive cellulose or gum.Tensioactive celluloses and gums can also act to emulsify and pullparticles and essential oils into solution. In certain embodiments,additional surfactants, which may have a harsh effect on cells, are notneeded or included in the topical, isotonic compositions. In certainembodiments, the viscosity-increasing agent comprises guar gum,methylcellulose, ethylcellulose, ethyl methyl cellulose, ethylhydroxyethyl cellulose, carboxymethylcellulose, hydroxypropylcellulose,hydroxyethyl methyl cellulose, hydroxypropylmethylcellulose(hypromellose), hydroxyethylcellulose, cetyl hydroxyethycellulose,hydroxypropyl guar gum glycosaminoglycans (e.g., hyaluronic acid),nonionic triblock copolymers such as poloxamers, gelatins, alginates,carrageenan, and agar, or any combination thereof. In some embodiments,the compositions may comprise a viscosity-increasing agent comprisingglycosaminoglycans (e.g., hyaluronic acid), nonionic triblock copolymerssuch as poloxamers, gelatins, carrageenan, agar, and combinationsthereof.

In certain embodiments, the topical, isotonic composition may furthercomprise a pH modifying agent to adjust the final pH of the compositionto the target or desired pH. The pH modifiying agent may comprise anacidifying agent, an alkalinizing agent, and/or both an acidifying agentand an alkalinizing agent. In certain embodiments, the pH modifyingagent is in an amount ranging from about 0.01% to about 1%. Exemplaryacidifying agents include, but are not limited to, organic acids, suchas citric acid, lactic acid, formic acid, acetic acid, propionic acid,butyric acid, caproic acid, oxalic acid, maleic acid, benzoic acid,carbonic acid, and the like. Exemplary alkalinizing agents includeammonia, ammonium carbonate, diethanolamine, monoethanolamine, potassiumhydroxide, potassium phosphate dibasic, sodium bicarbonate, sodiumborate, sodium carbonate, sodium hydroxide, sodium lactate, sodiumphosphate dibasic, trolamine, or any combination thereof. In certainembodiments, the pH of the topical, isotonic composition is formulatedat a pH to match to a normal, physiological fluid pH (e.g.,cervo-vaginal secretions, semen) or the epithelial surface of thegenital tissue, or the anogenital epithelium (e.g., mucosa, skin). Incertain embodiments, the pH of the topical, isotonic composition is lessthan about 6.8 or less than about 6.5 or less than about 6.2 or lessthan about 6.0 or less than about 5.7 or less than about 5.5. In certainembodiments, the pH of the topical isotonic composition is about 3.5 toabout 7.5, about 3.5 to about 6.8, about 3.5 to about 6.0, about 3.5 toabout 5.7, or about 6 to about 8. In a particular embodiment, the pH ofthe topical, isotonic composition is about is about 4.5, about 5, about6.5, or about 6.8. Topical, isotonic compositions having a pH of about3.5 to about 6, are particularly suited for administration to a subjectthat is aged about from adolescence to menopause/andropause, e.g., about18 years of age to about 50 years of age, or a child at least one yearold. In certain embodiments, an isotonic, topical composition foradministration to a subject of more than about 12 years of age (e.g.,from about 12 years of age to about 50 years of age, more than about 50years of age, more than 60 years of age, more than 70 years of age, morethan 80 years age, more than 90 years of age, more than 100 years ofage) has a pH in the urogenital and/or anogenital region of about 4.5 orabout 4.8. In certain embodiments, an isotonic, topical composition foradministration to a child at least one year old has a pH in theanogenital and/or urogenital region of about 5. Topical, isotoniccompositions having a pH of about 5 to about 7 are particularly suitedfor administration to an infant aged less than about 12 months old, to asubject of about 18 years of age to about 50 years of age who is tryingto conceive a child, or a senior of at least about 60 years of age. Incertain embodiments, a topical, isotonic composition for administrationto an infant has a pH in the anogenital and/or urogenital region ofabout 6.5. In certain embodiments, a topical, isotonic composition foradministration to a subject of about 18 years of age to about 50 yearsof age who is trying to conceive a child has a pH in the anogenitaland/or urogenital region of about 6.8.

In certain embodiments, topical, isotonic compositions of the presentdisclosure further comprise a buffering agent. The buffering agent maybe in an amount ranging from about 0.01% to about 0.9% by weight of thecomposition. A buffering agent refers to a compound or a mix ofcompounds that, when present in a solution, resists changes in the pH ofthe solution when small quantities of acid or base are added or upondilution with a solvent or bodily fluid. Buffer capacity is a measure ofthe resistance to change in the pH of a solution when acids or bases areadded to the solution. The total amount of the buffering agent (e.g.,conjugate acid-base pair) is selected such that the pH of thecomposition is maintained at the desired pH or range of pH values. Thus,the greater the amount of the buffering capacity, the more resistant thepH of the composition is to change. In certain embodiments, a bufferingagent contains an acidic species to neutralize hydroxide (OH⁻) ions anda basic species to neutralize hydrogen (H⁺) ions. However, the acidicand basic species of the buffering agent should not consume each otherthrough a neutralization reaction.

In certain embodiments, the buffering agent is a simple bufferedsolution comprising a weak acid and a salt of the weak acid or a weakbase and a salt of the weak base. Thus, the buffering agent can includea weak acid-base conjugate pair or weak base-acid conjugate pair.Examples of weak acid/salt of weak acid and weak base/salt of weak baseparings include citric acid/sodium citrate, lactic acid/sodium lactate,acetic acid/sodium acetate, monosodium phosphate/disodium phosphate,propionic acid/sodium propionate, butyric acid/sodium butyrate, carbonicacid/sodium bicarbonate, malic acid/sodium malate, ascorbic acid/sodiumascorbate benzoic acid/sodium benzoate, succinic acid/sodium succinateand sodium borate/boric acid. In certain embodiments, the bufferingagent comprises unrelated weak acid-base pairs. Examples of suchcombinations include disodium phosphate/citric acid, disodiumphosphate/lactic acid, monosodium phosphate/sodium lactate, monosodiumphosphate/sodium citrate, sodium citrate/lactic acid, sodiumlactate/citric acid, monopotassium phosphate/citric acid, monopotassiumphosphate/lactic acid, monopotassium phosphate/sodium lactate,monopotassium phosphate/sodium citrate, monopotassium citrate/lacticacid, and potassium lactate/citric acid. In addition, for multivalentanions, the calcium salt rather than sodium salt may be used (e.g.,calcium citrate). Example buffer can also includegluconolactone/gluconic acid.

In certain embodiments, the buffering agent is selected such that thebuffering agent's acid form has a pKa the same as or close to thedesired pH of the composition or a pH within the desired range of pHvalues, functions with a similar buffering capacity to surfaces andfluids that physiologically occur in genital region (e.g., vaginalmucin-acidic barrier, cervico-vaginal secretions, semen, menses-flow, ora combination thereof), or maintains pH to that of the target epithelialsurface (Rastogi et al., Contraception. 93:337, 2016). In certainembodiments, a buffering agent comprises a monocarboxylate, adicarboxylate, a carboxylic acid, or a combination thereof. In someembodiments, a buffering agent may comprise an acetate, borate, citrate,fumarate, lactate, malate, malonate, nitrate, phosphate, propanoate,succinate, tartrate, tromethamine, or any combination thereof. In someembodiments, a buffering agent comprises lactic acid, sodium lactate,sodium phosphate (monobasic, dibasic, or both), potassium phosphate(monobasic, dibasic, or both), sodium citrate, potassium citrate,calcium citrate, acetic acid, sodium acetate, citric acid, disodiumcitrate, trisodium citrate, boric acid/sodium, succinic acid, sodiumsuccinate, gluconolactone, disodium succinate, tartaric acid, sodiumtartarate, sodium ascorbate, ascorbic acid, tromethamine (Tris), or anycombination thereof. In certain embodiments, a buffering agent comprisescitric acid and disodium phosphate, lactic acid and sodium lactate,gluconolactone, or mono- or disodium phosphate and lactic acid.

Topical compositions of the present disclosure are specifically isotonicto the target genital fluids or tissues that they will contact. Tonicityis a measure of the effective osmotic pressure gradient (as defined bythe water potential of two solutions) of two solutions separated by asemipermeable membrane. Tonicity is commonly used when describing theresponse of cells immersed in an external solution. In other words,tonicity is the relative concentration of solutions that determine thedirection and extent of diffusion from a fluid across cell membranes intissue. Blood normally has an osmotic pressure that corresponds to thatof a 0.9% solution of sodium chloride (˜280 mOsm/kg). However, osmoticpressure for physiologic fluids occurring in the genital region can varywidely, from the low end for cervico-vaginal fluids (e.g., ˜128mOsm/kg), to ˜280 mOsm/kg for menses blood, to a higher level for semen(e.g., ˜320+ mOsm/kg). A composition (e.g., solution or gel) isconsidered isotonic when its tonicity matches that of the physiologicfluids it will contact. A composition is isotonic with a body fluid whenthe magnitude of the salts (ions) are equal between the composition andthe physiologic fluid. Tonicity equilibrium is reached in physiologicfluids by water moving across cell membranes, but the salts and ionsstaying in their fluid of origin. A solution is isotonic with a livingcell if there is no net gain or loss of water by the cell, or otherchanges in the cell ultrastructure, when it is in contact with saidsolution, even though individual water molecules may move freely acrossthe cell membranes.

Hypertonic solutions cause a net movement of water out of the cells (asthe water moves to create equilibrium with the high salt levels outsideof the cell). This dehydration of the cell is concentration dependentand leads to osmotic stress which can increase reactive oxygen species,cause cytoskeletal rearrangement, and damage DNA and mitochondrialfunction within minutes of exposure. Most current genital products arehypertonic, resulting in epithelial cell and sperm death on contact.Hypotonic solutions cause a net flow of water into the cell and causecell bursting and death. Some deviations of salt levels in physiologicfluids from the level found in blood and tissues may serve a purpose.For example, the lower osmolality of cervico-vaginal fluids thatfacilitates vaginal epithelial cell lysis and death as a part of normalvaginal function. In another example, the higher osmolality of semen canprotect sperm cells from the lower osmolality of cervico-vaginalsecretions following ejaculation in the vagina and admixing of fluidsduring vaginal intercourse.

Related to tonicity is osmosis, which is the movement of solvent acrossa semipermeable membrane from an area of higher solute concentration toan area of lower solute concentration to produce equilibrium. Osmoticpressure of a solution is the pressure that must be applied to stop theflow of solvent across a semipermeable membrane.

In certain embodiments, the compositions of the present disclosurefurther comprise an osmolality adjusting agent to adjust the tonicity ofthe compositions. Exemplary osmolality adjusting agents includeelectrolytes, mono- or disaccharides, inorganic salts (e.g., sodiumchloride, calcium chloride, potassium chloride, sodium sulfate,magnesium chloride), or a combination thereof. In some embodiments, anosmolality adjuster is glucose, sucrose, sodium chloride, potassiumchloride, calcium chloride, sodium sulfate, magnesium chloride,dextrose, mannitol, or any combination thereof.

In certain embodiments, the osmolality range of the compositionsdisclosed herein ranges from about 120 mOsm/kg to about 450 mOsm/kg orfrom about 240 mOsm/kg to about 450 mOsm/kg. In certain embodiments, theosmolality of the compositions of the present disclosure is about 120mOsm/kg, about 125 mOsm/kg, about 130 mOsm/kg, about 135 mOsm/kg, about140 mOsm/kg, about 145 mOsm/kg, about 150 mOsm/kg, about 155 mOsm/kg,about 160 mOsm/kg, about 165 mOsm/kg, about 170 mOsm/kg, about 175mOsm/kg, about 180 mOsm/kg, about 185 mOsm/kg, about 190 mOsm/kg, about195 mOsm/kg, about 200 mOsm/kg, about 205 mOsm/kg, about 210 mOsm/kg,about 215 mOsm/kg, about 220 mOsm/kg, about 225 mOsm/kg, about 230mOsm/kg, about 235 mOsm/kg, about 240 mOsm/kg, about 245 mOsm/kg, about250 mOsm/kg, about 255 mOsm/kg, about 260 mOsm/kg, about 265 mOsm/kg,about 270 mOsm/kg, about 280 mOsm/kg, about 285 mOsm/kg, about 290mOsm/kg, about 295 mOsm/kg, about 300 mOsm/kg, about 305 mOsm/kg, about310 mOsm/kg, about 315 mOsm/kg, about 320 mOsm/kg, about 325 mOsm/kg,about 330 mOsm/kg, about 335 mOsm/kg, about 340 mOsm/kg, about 345mOsm/kg, about 350 mOsm/kg, about 355 mOsm/kg, about 360 mOsm/kg, about365 mOsm/kg, about 370 mOsm/kg, about 375 mOsm/kg, about 380 mOsm/kg,about 385 mOsm/kg, about 390 mOsm/kg, about 395 mOsm/kg, about 400mOsm/kg, or about 450 mOsm/kg.

In certain embodiments, the topical, isotonic composition is matched fortonicity (e.g., salt/ion levels) to the normal, physiological genitalfluid pH (e.g., CVF, urethral secretions, semen, smegma) of the subject;genital tissue of the subject (e.g., vaginal mucosa, genital skin); orat an appropriate tonicity for the particular method of use (e.g., foruse during fertile window in a subject or sexual dyad that is trying toconceive). In certain embodiments, the tonicity ranges from about 125mOsm/kg to about 240 mOsmo/kg. Such embodiments match the hypotonic CVFwhich supports lysis of vaginal epithelial cells and vaginal“self-cleaning.” This cell lysis releases glycogen, which healthygenital microbiota utilize for growth and development. Such embodimentsare ideal for delivery inside the vaginal canal. In other embodimentsthe tonicity ranges from about 240 mOsm/kg to about 280 mOsm/kg. Suchembodiments match the tonicity of genital tissues and are ideal forcontact with skin genital tissue surfaces. In a particular embodiment,the tonicity ranges from about 280 mOsmo/kg to about 450 mOsmo/kg tomatch the tonicity of semen as deposited in the vagina. In certainembodiments, tonicity may be expressed as mOsm/kg or mOsm/L. Osmolalityis a measure of the osmoles (Osm) of solute per kilogram of solvent.Depending on the density of the solvent, the osmolality and osmolarity(Osm of solute per L of solution) may differ. In certain embodiments,the osmolality and osmolarity values are substantially interchangeable.

In certain embodiments, the topical, isotonic composition furthercomprises a solvent (e.g., aqueous solvent, water) in an amount greaterthan about 88% (e.g., ranging from about 88% to about 98%). In furtherembodiments, the solvent comprises water.

In certain embodiments, the topical, isotonic composition furthercomprises an additional therapeutic agent. The additional therapeuticagent may improve cell or tissue function or treat an underlying diseaseor disorder. In certain embodiments, the therapeutic agent is anantibiotic, anti-fungal agent, anti-viral agent, or any combinationthereof. Exemplary anti-fungal agents include butoconazole nitrate,clotrimazole, miconazole nitrate, terconazole, tioconazole, econazolenitrate, efinaconazole, ketoconazole, luliconazole, naftifinehydrochloride, oxiconazole nitrate, sertaconazole nitrate, sulconazolenitrate, tavaborole, terbinafine, acyclovir, tenovir, zidovudine,stavudine, metronidazole, or a combination thereof. In some embodiments,the additional therapeutic agent is a vaccine (e.g., multivalentvaccine) to provide immunity against a viral or bacterial disease.Suitable vaccines include uropathogenic Escheri coli bacteria asdisclosed in W Hopkins, et al., J Urol., 177 (2007): 1349-1353, herebyincorporated by reference in its entirety, and particularly in relationto the vaccine suppositories used in the study. In some embodiments, thevaccine may include a cholera vaccine such as those as disclosed in PKozlowski, et al., Infection and Immunity 65 (1997): 1387-1394, which ishereby incorporated by reference in its entirety and particularly inrelation to cholera vaccines, and vaccines containing killed Vibriocholerae cells. In some embodiments, the additional therapeutic agentmay treat or prevent atrophic vaginitis. Suitable agents for thetreatment or prophylaxis of atrophic vaginitis include hyaluronic acid,estrogens including estradiol-17β, conjugated estrogens, estradiolhemihydrate, dehydroepiandrosterone, estradiol acetate, selectiveestrogen receptor modulators, including bazedoxifene, cyclofenil,lasofoxifene, ormeloxifene, ospemifene, raloxifene, toremifene, andcombinations thereof.

In certain embodiments, the additional therapeutic agent is a topicalpain relieving agent. Exemplary topical pain relieving agents includelidocaine, benzocaine, novocaine, diphenhydramine, and pramoxine.

Other examples of therapeutic agents include hormones (e.g., estradiol,estriol, estropipate, testosterone, progesterone, DHEA, testosterone, ora combination thereof), contraceptive agents (e.g., impairs spermfunction, thickens cervical mucus, or both), agents that enhancevasodilation (e.g., In some embodiments, an agent that enhancesvasodilation is L-arginine, niacin, nicotinamide, alprostadil, aphosphodiesterase inhibitor), erectile dysfunction treatment or erectileenhancement drugs (e.g., alprostadil, glyceryl trinitrate, sildenafil,tadalafil, vardenafil, avanafil, lodenafil, mirodenafil, udenafil,zaprinast, Icariin, benzamidenafil, dasantafil), and prematureejaculation drugs (e.g., selective serotonin reuptake inhibitorsincluding sluoxetine, paroxetine, sertraline; tricyclic antidepressantsincluding clomipramine).

Yet another example of a therapeutic agent is a skin conditioner oremollient.

In certain embodiments, the topical, isotonic compositions of thepresent disclosure further comprise at least one genital probioticbacterial species or strain (e.g., belonging to the genusLactobacillus). In certain embodiments, the probiotic bacterial speciesor strain is one having the ability to colonize the human vagina orpenis. The adhesion of lactobacilli to the uroepithelium varies amongspecies and strains, as shown by in vitro studies (Reid et al., J. Urol.138:330, 1987), and may be mediated by glycoprotein and carbohydrateadhesins binding to glycolipid receptors (Boris et al., Infection andImmunity 66:1985, 1998). In some embodiments, a genital probioticspecies is a species that is part of the genital microbiota (e.g.,vagina or penis). In a specific embodiment, a genital probiotic speciesis selected from Lactobacillus acidophilus, Lactobacillus jensenii,Lactobacillus gasseri, Lactobacillus iners, Lactobacillus crispatus,Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus brevis,Lactobacillus casei, Lactobacillus delbrueckii, Lactobacillus vaginalis,Lactobacillus salivarius, Lactobaccillus reuteri, and Lactobacillusrhamnos, Streptococcus, non-pathogenic Prevotella species, Bacillus, orany combination thereof.

In certain embodiments, the topical, isotonic compositions of thepresent disclosure may further comprise a preservative. The preservativemay be in an amount of about 0.001% to about 4% (e.g., 0.001% to about1%) by weight of the composition. Exemplary preservatives includecaprylyl glycol, cranberry extract, dichlorobenzyl alcohol,gluconolactone, green tea extract, oleuropein, pentylene glycol,phenethyl alcohol, pomegranate extract, potassium benzoate, propanediol,resveratrol, hydantoin, benzoic acid, benzyl alcohol, dihydroaceticacid, ethylhexyl glycerin, Lactobacillus ferment, pentylene glycol,potassium sorbate, sodium benzoate, sodium dehydroacetate, sodiumsalicylate, or any combination thereof.

In certain embodiments, the topical, isotonic compositions of thepresent disclosure are sterile and/or preservative-free.

The topical, isotonic compositions of the present disclosure may furthercomprise additional pharmaceutical excipients. Pharmaceuticallyacceptable excipients for therapeutic use are well known in thepharmaceutical art, and are described herein and, for example, inRemington's Pharmaceutical Sciences, Mack Publishing Co. (A. R. Gennaro,ed., 18^(th) Edition, 1990) and in CRC Handbook of Food, Drug, andCosmetic Excipients, CRC Press LLC (S.C. Smolinski, ed., 1992).

Compositions of the present disclosure can be formulated as a liquid,semi-solid, soap, gel, jelly, film, foam, cream, douche, ointment,lotion, spray, aerosol, suspension, emulsion, or paste. In certainembodiments, the topical, isotonic compositions are formulated in asingle-use or unit-dose format.

In certain embodiments, topical, isotonic compositions of the presentdisclosure are integrated into a tampon, vaginal ring, cervical cup,diaphragm, condom, wipe, blanket, undergarment, or diaper. In certainembodiments, the topic, isotonic compositions are administered using asyringe, a roller ball, foam dispenser, spray bottle, aerosol dispenser,or pump dispenser.

In certain embodiments, compositions of the present disclosure areintegrated into a microbiota sample collection and recovery system, suchas a sterile swab or cyto-brush. In certain embodiments compositions ofthe present disclosure are used to facilitate the transplantation,storage, and cultivation of desirable gastrointestinal, vaginal,genital, reproductive tract, urinary tract, and/or anogenital microbiotafrom healthy donors for use in the treatment of microbial dysbiosis inaffected recipients. For example, a vaginal rinse, using thecompositions of the present disclosure may be collected from the vaginaof healthy user and transferred to a subject with a dysbiotic microbiomeof the anogenital and/or urogenital region, to promote beneficialmicrobiome transplantation. Such procedures are described in ALev-Sagie, et al., Nature Medicine 25 (2019): 1500-1504, herebyincorporated by reference in its entirety. In another example, using thecompositions of the present disclosure, a wipe may be used by a parentto collect beneficial genital microbiome species to transfer these viatopical application to an infant born by Cesarean section. In certainembodiments, the composition is tailored to the individual recipient anddonor profile to maximize the efficacy of the microbiota transplantationfor each individual (e.g., the composition may be formulated for theproduction of specific bacteria in a user's microbiome). In certainembodiments, the composition would sustain beneficial microorganismssuch as Lactobacillus acidophilus, Lactobacillus jensenii, Lactobacillusgasseri, Lactobacillus crispatus, Lactobacillus plantarum, Lactobacillusfermentum, Lactobacillus brevis, Lactobacillus casei, Lactobacillusdelbrueckii, Lactobacillus vaginalis, Lactobacillus salivarius,Lactobaccillus reuteri, Lactobacillus rhamnosus, and combinationsthereof. In addition compositions may also sustain other bacterialspecies of the Leptotrichia, Leuconostoc, Pediococcus, Akkermansia,Streptococcus, Faecalibacterium and Weissella genera during the timeinterval between collection and implantation. In certain embodiments,compositions of the present disclosure may inhibit and/or minimize thegrowth of organisms that are or can become pathogenic including thespecies Lactobacillus iners, or species from the genus Prevotella,Eggerthellia, Gardnerella, Atopobium, Megasphaera, Mobiluncus,Mageeibacillus, Gemella, Veillonella Sneathia Clostridium, andcombinations thereof. The growth inhibition and/or minimization mayoccur in microbial transplantation sample storage media.

In certain embodiments, compositions of the present disclosure may beused to add and facilitate the implantation process associated withtampons, and/or reusable silicon devices such as menstrual cups for thecontrol of natural menstrual flow in women or lochia post-partum. Kitsare also provided comprising a device for insertion into the vagina forcontrol of natural menstrual flow such as a tampon and/or these reusablesilicon device, wherein the device is packaged in a composition of thepresent disclosure. For example, the compositions may be formulated witha pH-balanced gel for use in a tampon lubricant. These gels may containa lactic acid buffer designed to maintain vaginal pH between 3.8 and 4.2for about 5 to about 10 hours or about 6 to about 9 hours or about 8hours.

The compositions may be formulated with thermoresponsive polymers whichundergo a phase change exhibiting a sol-gel transition in response tobody temperature, pH, and specific ions present in physiologicalenvironments. Such thermoresponsive polymers may prolong the residencetime of the composition in the urogenital and/or anogeital region (e.g.,vagina). Exemplary thermoresponsive polymers include poloxamers,styrene-butadiene block copolymer, polymethylacrylate,polybutylmethacrylate, plasticized polyvinylchloride, plasticized nylon,plasticized polyethyleneterephthalate, polyethylene, polyacrylonitrile,polytrifluoro chloroethylene, poly-4,4′-i sopropylenediphenylenecarbonate, polyethylenevinyl esters, polyvinylchloride-diethyl fumarate,and combinations thereof.

In various implementations, the compositions further comprise one ormore sustained release polymers. Suitable sustained release polymersinclude sodium alginate (e.g., with barium chloride), barium chloride,poly-l-lysine, polyvinylamine, protamine sulfate, and combinationsthereof.

Vaginal rings or pesseries are devices inserted in the vagina to achievecontrolled release of the active composition. Medicated vaginal ringsmay be fabricated from Silastic® 382 medical grade elastomer. The mostcommonly used polymers for vaginal ring are ploy (dimethylsiloxane) orsilicon devices, or ethylene vinyl acetate. Additoinally, biodegradablepolymers, such as polycaprolactone have been used to prepare thesedevices. These are generally polymeric rings in which the drug (e.g.,borneol, pharmaceutically acceptable salts thereof, or prodrugs of anyof the foregoing such as bornyl acetate, prebiotic oligosaccharide,metal co-factor) and/or composition is homogeneously dispersed. In orderto achieve constant release, two types of system are typically developedfor vaginal rings: sandwich and reservoir type. In the sandwich type, anarrow layer of the active components is placed between non-medicatedcentral core and non-medicated outer band. In reservoir type, centralcore having the active components is encapsulated with drug-free polymerlayer (e.g., hypromellose).

The compositions and/or active ingredients (e.g., borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing such as bornyl acetate, prebiotic oligosaccharide, metalco-factor) may also be formulated in vaginal suppositories, ovules, orpessaries. In these embodiments, the suppository, ovule, or pessary maybe formulated with mixture of synthetic triglycerides (e.g. WitepsolH-15), hypromellose, polyethylene glycol polymers (e.g. PEG 1000, 4000),Sorbitane monostearate (Span 60), PEG-35 Castor Oil, PEG 400, PEG 3350,cocoa butter, polyethylenimine, mixtures of mono/di and triglycerides(e.g. Suppocire, Ovucire), Agar, Propylene glycol, Hypromellose,gelatin, glycerin, cocoa butter, bees wax, Polyoxyl 40 stearate,Polyvinyl alcohol, hydroxyethyl methacrylate, polyacrylic acid,polyoxyethylene, and combinations thereof.

In some embodiments, the composition may be formulated as an emulsion.The emulsion may be, for example, a water-in-oil, oil-in-water,silicone-in-water, water-in-silicone, polyol-in-oil, oil-in-polyol,glycerin-in-oil, oil-in-glycerin, silicone-in-glycerin,glycerin-in-silicone, silicone-in-polyol, or polyol-in-siliconeemulsion. The topical isotonic formulations of the present disclosuremay be used as the aqueous phase (e.g., the internal phase, thediscontinuous phase) of an emulsion. In an emulsion, will be understoodthat the weight percentage of components used herein (e.g., borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing such as bornyl acetate, prebiotic oligosaccharide, metalco-factor) refers to the weight percentage of the composition (and notthe weight percentage of the phase itself). The nonaqueous phase maycomprise saturated (lauric, myristic and capric acid) and unsaturatedfatty acids (oleic acid, linoleic acid and linolenic acid), surfactants,and co-surfactants. Exemplary components in emulsions include sorbitanoleate (Span80), Sorbitane trioleate (Span85), polyethylene glycolsorbitan monolaurate (Tween80), Soybean oil, squalene, lecithin, oleicacid, medium chain triglyceride, Polyoxyl 40 Hydrogenated Castor Oil,Polyoxyl 35 castor oil, Glycerol, Propylene glycol, and combinationsthereof.

In certain implementations, the emulsion is a Pickering emulsion.Typically Pickering emulsions are emulsions stabilized by solidparticles including nanocellulose, graphene oxide, carbon nanotube,carbon lamp black, laponite, montmorillonite, silica nanoparticles,calcium carbonate, titanium dioxide, magnetic particles, polymerparticles, and combinations thereof.

In some embodiments, the compositions or active ingredients (e.g.,borneol, pharmaceutically acceptable salts thereof, or prodrugs of anyof the foregoing such as bornyl acetate, prebiotic oligosaccharide,metal co-factor) are formulated as a tablet. For example, thecomposition may be in tablet form such as chitosan and/or sodiumalginate based bio-adhesive tablets. In certain embodiments, the tablefurther comprises a mucoadhesive. The compositions or active ingredients(e.g., borneol, pharmaceutically acceptable salts thereof, or prodrugsof any of the foregoing such as bornyl acetate, prebioticoligosaccharide, metal co-factor) may be formulated as vaginalbioadhesive tablets. Vaginal bioadhesive tablets may comprisehydroxypropyl cellulose, polyacrylic acid, Carbopol-934, andcombinations thereof.

In some embodiments, the compositions or active ingredients (e.g.,borneol, pharmaceutically acceptable salts thereof, or prodrugs of anyof the foregoing such as bornyl acetate, prebiotic oligosaccharide,metal co-factor) are formulated as liposomes (e.g., vaginal liposomes).Vaginal liposomes include lecithin based liposomes which may incorporatebio-adhesive carbopol hydrogels. In some embodiments, the compositioncomprises a thermo-sensitive gel of poloxamers 407 and 188 and are inthe form of active ingredients (e.g., borneol, pharmaceuticallyacceptable salts thereof, or prodrugs of any of the foregoing such asbornyl acetate, prebiotic oligosaccharide, metal co-factor) loadedcationic liposomes.

The compositions (e.g., gels), applicators, and/or kits of the presentdisclosure may be subjected to ozone sterilization. Wherein embodimentsof the present disclosure may be exposed to ozone gas due to itsoxidative potential. The possibility to alter different processparameters (e.g., time of exposure, gas concentration, humidity) allowsthe sterilization protocol to be adapted to different types of material.

Exemplary formulations according to the present disclosure are providedin Tables 1-7.

TABLE 1 Lubricant, pH 4.5, osmolality 150 mOsm/kg Ingredient Name % bywt. Purified water, USP 96.1422 Disodium phosphate 0.39825 Lactic Acid0.1425 Lactulose 0.05 Abies sibirica (siberian fir) 0.02 hydroxyethylcellulose 0.8 Oleuropein 0.02 gluconolactone 1.00 hydroxypropyl guar gum0.4 Arabinogalactan 1.00 Mentha spicata 0.0075 Citrus reticulata 0.0075Juniperus communis 0.0075 Manganese chloride* 0.0046 *The indicatedweight percentage of metallic co-factor (i.e., magnesium chloride) maybe achieved by adding the appropriate amount of a hydrate of themetallic salt (e.g., magnesium chloride (II) tetrahydrate). For example,in this embodiment the composition may comprise 0.0046% (w/w) MnCl₂ (MW= 125.9 g/mol) as added from MnCl₂•4H₂O (MW = 197.9 g/mol). Therefore,0.00723 g hydrate were added per 100 g of formulation.

TABLE 2 Foaming Gel, pH 4.5, osmolality 120 mOsm/kg Ingredient Name % bywt. Purified Water, USP 93.840 Manganese chloride* 0.008 Lactulose 0.05Abies sibirica (siberian fir) 0.02 Cetyl Hydroxyethylcellulose 0.5Oleuropein 0.02 sodium cocoyl glutamate 2.00 Sodium Lauroamphoacetate2.00 Arabinogalactan 1.00 disodium phosphate 0.39825 Lactic Acid 0.1425Mentha spicata 0.008 Citrus reticulata 0.005 Juniperus communis 0.008*The indicated weight percentage of metallic co-factor (i.e., magnesiumchloride) may be achieved by adding the appropriate amount of a hydrateof the metallic salt (e.g., magnesium chloride (II) tetrahydrate). † Theindicated weight percent of Cocamidopropyl betaine is the weight percentfrom a 36% solutions of cocamidopropyl betaine may be added to eachsolution.

TABLE 3 Wipe Formula, pH 6.5 (infant 0-12 mo) or pH 5 (child >1 yr), 180mOsm/kg Ingredient Name % by wt. Purified Water, USP 94.861 Lactulose0.100 Abies sibirica (siberian fir) 0.020 Cetyl Hydroxyethylcellulose0.500 Oleuropein 0.020 Cocamidopropyl Betaine^(†) 2.100 Arabinogalactan0.900 Citric Acid 0.098 disodium phosphate 0.386 Citrus reticulata 0.010Manganese chloride* 0.0046 sodium benzoate 0.3000 gluconolactone 0.7000*The indicated weight percentage of metallic co-factor (i.e., magnesiumchloride) may be achieved by adding the appropriate amount of a hydrateof the metallic salt (e.g., magnesium chloride (II) tetrahydrate).^(†)The indicated weight percent of Cocamidopropyl betaine is the weightpercent from a 36% solution of cocamidopropyl betaine may be added toeach solution.

TABLE 4 Lubricant, pH 6.8, osmolality 340 mOsm/kg Ingredient Name % bywt. Purified Water, USP 95.67 disodium phosphate 0.70 Lactic Acid 0.18Lactulose 0.03 sodium chloride 0.15 Raffinose 0.02 Abies alba 0.01hydroxyethyl cellulose 0.80 Oleuropein 0.02 Hydantoin 1.00 hydroxypropylguar gum 0.40 Arabinogalactan 1.00 Monarda Fistulosa 0.01 Citrusreticulata 0.01 Rosmarinus officinalis 0.00 Manganese chloride* 0.01*The indicated weight percentage of metallic co-factor (i.e., magnesiumchloride) may be achieved by adding the appropriate amount of a hydrateof the metallic salt (e.g., magnesium chloride (II) tetrahydrate).

TABLE 5 Douche formulation, pH 4.5 Ingredient Name % by wt. PurifiedWater, USP 98.019 Monosodium Phosphate, anhydrous 0.483 Sodium Chloride0.100 Lactulose 0.050 Abies sibirica (Siberian fir) 0.020 Hypromellose0.600 Gluconolactone 0.250 Hydroxypropyl guar gum 0.400 Arabinogalactan0.050 Mentha spicata (spearmint) 0.015 Neroli oil 0.010 Manganesechloride* 0.003 Lactic Acid 10% Titrate to pH 4.5 *The indicated weightpercentage of metallic co-factor (i.e., magnesium chloride) may beachieved by adding the appropriate amount of a hydrate of the metallicsalt (e.g., magnesium chloride (II) tetrahydrate).

TABLE 6 Douche formulation, pH 4.5 Ingredient Name % by wt. PurifiedWater, USP 99.061 Monosodium Phosphate, anhydrous 0.241 Sodium Chloride0.200 Lactulose 0.050 Abies sibirica (Siberian fir) 0.020 hypromellose0.400 Mentha spicata (spearmint) 0.015 Neroli Oil 0.010 Manganesechloride* 0.003 Lactic acid 10% Titrate To pH 4.5 *The indicated weightpercentage of metallic co-factor (i.e., magnesium chloride) may beachieved by adding the appropriate amount of a hydrate of the metallicsalt (e.g., magnesium chloride (II) tetrahydrate).

TABLE 7 Foaming Gel formulation, pH 4.6 Ingredient Name % by wt.Purified Water, USP 97.3375 Sodium chloride 0.1000 Lactulose 0.0500Rosemary essential oil 0.0500 Abies sibirica (Siberian fir) 0.0100Hypromellose 0.3000 Green tea polyphenols 0.0200 Arabinogalactan 0.1000Mentha spicata (spearmint) 0.0200 Neroli Oil 0.0100 Manganese chloride0.0025 Sodium Cocyl Isethionate 1.0000 Cocamidopropyl Betaine† 1.0000*The indicated weight percentage of metallic co-factor (i.e., magnesiumchloride) may be achieved by adding the appropriate amount of a hydrateof the metallic salt (e.g., magnesium chloride (II) tetrahydrate). †Theindicated weight percent of Cocamidopropyl betaine is the weight percentfrom a 36% solution of cocamidopropyl betaine may be added to eachsolution.

It will be understood that components may have multiple purposes asthose described herein. For example, citric acid may be considered a pHmodifying agent and a buffering agent.

In Vitro and In Vivo Activity

(1) Evaluating Genital Microbiota

To assess whether topical, isotonic compositions of the presentdisclosure are harmful to the genital microbiota, testing of normalgenital microbiota may be performed using methods known in the art.

The effect of any composition disclosed herein on genital microbiota(e.g., Lactobacillus species) may be determined by measuring minimalinhibitory concentration, the lowest concentration which preventsvisible growth of a microorganism after overnight culture, or minimalmicrobicidal concentration, the lowest concentration required to reducethe viability of a culture by 99.99%.

In some embodiments, the effect of any composition disclosed herein ongenital microbiota (e.g., Lactobacillus species) may be determined byDNA extraction and 16S rRNA gene sequencing and operational taxonomicunits (OTUs) assignment and community states (CST) of vaginal microbiomecan be defined using Jensen-Shannon divergence and Ward linkagehierarchical clustering following administration, as disclosed in XHong, et al., PeerJ 7 (2019): e8131, which is hereby incorporated byreference in its entirety.

The following signs and symptoms are all associated with genitalmicrobiome function and may be used to assay effectiveness of theproducts. Vaginal infection is highly correlated with Lactobacillusdominance of the genital microbiome. Infection may be determined byexamining a wet mount smear in potassium hydroxide for detection ofcandidiasis and in saline for detection of motile trichomonas and cluecells. Bacterial vaginosis (BV) is associated with Lactobacillusdominance as well. Women with BV have dysbiosis of the genitalmicrobiome. BV diagnosis may be assessed according to Amsel clinicalcriteria or Nugent testing or H₂O₂ and leukocyte esterase levels.Typically, a healthy vaginal microbiome will have lower levels of BV orother infections. BV diagnostic tests include: Nugent score, Amselcriteria, clue cell presence, whiff test and newer diagnostic tests(Aptima Bacterial Vaginosis and Aptima Candida/Trichomonas VaginitisAssays). The ‘Amsel's criteria’ requires that three of the followingfour criteria be met for BV: first, a vaginal pH of greater than pH 4.5;second, the presence of clue cells in the vaginal fluid; third, a milky,homogeneous vaginal discharge; and finally, the release of an amine(fishy) odor after addition of 10% potassium hydroxide to the vaginalfluid. Suitable protocols for laboratory diagnosis of BV include thosedisclosed in D. Money. Can J Infect Dis Med Microbiol 16 (2005): 77-79,hereby incorporated by reference in its entirety, and specifically inrelation to BV diagnosis protocols.

Vaginal secretion measurements after use of the product may also bemeasured using a colorimetric or other assay for increased d-lactic acidand/or amylase as indicators of healthy microbiome as disclosed in JLeizer, et al., Reprod Sci. 25 (2018):854-860, or D Nasioudis et al.,Reprod. Sci 22 (2015): 1393-1398.

Vaginal Lactobacillus dominance has also been linked to high risk HPVinfection and persistence, genital herpes and HIV infections. MeasuringSTD levels in patients may help determine efficacy of the gel. Severalreports have shown that anti-HIV levels in CVF are higher in women withhealthy genital microbiomes as illustrated in M Torcia, Int J Mol Sci 20(2019): E266, S Placios et al., Minerva Ginecol 70 (2018): 138-143, andR Hemalatha et al., Indian J Med Res 138 (2013): 354-359, each of whichis hereby incorporated by reference in its entirety.

Vaginal pH may be measured by litmus paper, vaginal glove, electrode, ornanosensor as disclosed in U.S. Pat. No. 10,436,745, hereby incorporatedby reference in its entirety and particularly in relation to pHnanosensors. These measurements may be used to assess bothersome vaginalsymptoms and infection. For example, elevated vaginal pH is relativelysensitive for detecting BV and dysbiosis in women.

Vaginal bothersome symptom manifestation may be increased in womenwithout Lactobacillus dominance. Vaginal symptom level and sexualfunction can be tracked following product use using published tools asdisclosed in B Ettinger, et al., Menopause 5 (2008): 885-889, herebyincorporated by reference in its entirety. Furthermore, vaginalmetabolomics can be used to determine genital microbiome health, asdisclosed in T. Nelson, Front Physiol 6 (2015): 253, hereby incorporatedby reference in its entirety.

Biogenic amines and metabolites are also biomarkers of BV and dysbiosis,as they facilitate the outgrowth of BV-associated vaginal taxa by (i)amino-acid decarboxylation that consumes intracellular hydrogen ions andchange bacterial acid resistance; (ii) limiting the growth andresistance of host immunology to urogenital pathogens; and iii) beingcorrelated with numerous host disease states, including STDs, cancer anddementias. To measure, samples may be eluted from vaginal swabs insterile molecular water and subjected to both liquid and gaschromatography mass spectrometry.

The presence of pathogenic bacteria can also be assessed by measuringconcentration of endotoxins, LPS, and quantity of pathogenic bacteria invaginal washes obtained from subjects following the use of compositionsof the present disclosure (see, e.g., Aroutcheva et al., Anaerobe14:256, 2008). High LPS concentrations create a toxic vaginalenvironment causing epithelial and gamete (e.g. sperm) damage. Even verylow levels of LPS (e.g., 0.1 μ/mg) rapidly impact sperm function (see,Li et al., Tohoku Journal of Experimental Medicine 238:105, 2016,incorporated herein by reference in its entirety)

Other methods that may be used to assess changes in genital microbiotafollowing exposure to a topical, isotonic composition of this disclosureinclude performing 16S rRNA gene sequencing, shotgun metagenomic genesequencing, microbial/host metabolomic profiling, and 3^(rd) generationsequencing utilizing nanopore DNA sequencing (see, e.g., Romero et al.,Microbiome 2:4, 2014; Macklaim et al., Microb. Ecol. Health Dis.26:27799, 2015), species specific quantitative PCR (Zozaya-Hinchliffe etal., J. Clin. Microbiol. 48:1812, 2010, each of which is incorporatedherein by reference in its entirety), and phylogenetic microarrays(Paliy and Agans, FEMS Microbiol. Ecol. 79:2, 2012; Chen et al., NatCommun. 8:875, 2017, each of which is incorporated herein by referencein its entirety) using bacterial samples obtained from subjects (e.g.,washes, swabs).

In certain embodiments, the compositions of the present disclosure donot reduce normal genital microbiota population (e.g., L. crispatus, L.gassseri, L. jensenii, L. acidophilus, or any combination thereof) bymore than about 35%, about 30%, about 25%, about 24%, about 23%, about22%, about 21%, about 20%, about 19%, about 18%, about 17%, about 16%,about 15%, about 14%, about 13%, about 12%, about 11%, about 10%, about9%, about 8%, about 7%, about 6%, about 5%, about 4%, about 3%, about2%, or about 1% when exposed to a concentration or amount of thecomposition that is to be used in vivo on a subject.

In certain embodiments, the compositions of the present invention do notinterfere with acid-producing bacterial growth and functional mediumacidification of fluids or solutions of the genital microbiome (e.g.,Lactobacillus spp) (see, Boskey et al., Infect Immun. 67: 5170, 1999,incorporated herein by reference in its entirety).

In certain embodiments, the compositions of the present disclosure donot promote the growth of pathogenic bacteria of the anogenital regionby more than about 1%, about 2%, about 3%, about 4%, about 5%, about 6%,about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%,about 20%, about 25%, about 30%, or about 35%. In certain embodiments,the pathogenic bacteria is selected from pathogenic strains ofPrevotella, Eggerthellia, Gardnerella, Atopobium, Megasphaera,Mageeibacillus, Mobiluncus, Bacteroides, Peptostreptococcus,Fusobacterium, Veillonella, Porphyromonas and Eubacterium.

(2) Evaluating Genital Irritation, Inflammation, and Cell Death

Irritation, inflammation, or cell death of genital tissues, such as thevaginal mucosa or penile foreskin cells, can be assessed in vitro or invivo using human or animal vaginal-ectocervical, urethral, or foreskintissue explants; vaginal, cervical or vulvar cell monolayers; penileepithelium or urethral epithelium; skin cell monolayers; slug mucosalirritation assays; or other equivalent methods.

For example, the slug mucosal irritation assay (SMI) is a sensitivesystem to detect even mild mucosal irritation potency (Adriaens et al.,Sex. Transm. Dis. 35:512, 2008, incorporated herein by reference in itsentirety). The SMI assay uses slugs (Arion lusitanicus) as the testorganism. The body wall of slugs consists of a mucosal surfacecomprising mucus secreting cells covering a sub epithelial connectivetissue. Slugs that are placed on an irritant substance will activelyproduce mucus as a protective mechanism from noxious agents.Additionally, tissue damage of the slug's surface results in the releaseof proteins and enzymes. The protein concentration in the collectedsamples is determined with a protein quantitation kit. A composition ofthe present disclosure is considered non-irritating if it does not causean increased production of mucus, or an increased release of proteinsand enzymes as compared to a negative control.

Human, organotypic vaginal-ectocervical tissue models produced fromnormal human-derived vaginal-ectocervical epithelial cells may also beused to assess irritation of topically applied products, as canmonolayers of cervical or vaginal epithelium (Ayehunie et al.,Toxicology 279:130, 2011; Ayehunie et al., 2007, Toxicology 279:130,2007; Trifonova et al., Antimicrob. Agents Chemother. 50:4005, 2006;Fichorova et al., mBio 2:e00168, 2011, each of which is incorporatedherein by reference in its entirety). Release of markers of cell damage(e.g., increase in CD4, IL-1β, CXCL8, CCL2, CCL21, EMP1; decrease inBPI) and production of inflammatory mediators, such as IL-1, IL-8, TLR4,may be used as markers of irritation and pro-inflammation (see, also,Fichorova et al., Toxicol. Appl. Pharmacol. 285:198, 2015; Doncel etal., J. Acquir. Immune Defic. Syndr. 37(Suppl. 3):S174, 2004; Fichorovaet al., Biol. Reprod. 71:761, 2004; Moench et al., BMC Infect. Dis.10:331, 2010, each of which is incorporated herein by reference in itsentirety). Biomarkers of epithelial integrity and immune function havebeen validated in multiple clinical studies of vaginal product safety(Mauck et al., AIDS Res. Hum. Retroviruses 29:1475, 2013; Fichorova etal., mBio, 6: e00221, 2015; Fichorova et al., Cytokine 55:134, 2011;Mauck et al., J. Acquir. Immune Defic. Syndr. 49:243, 2008; Morrison etal., J. Acquir. Immune Defic. Syndr. 66:109, 2014; Schwartz et al.,Contraception 74:133, 2006; Keller et al., J. Antimicr. Chemother.51:1099, 2003, each of which is incorporated herein by reference in itsentirety). A composition of the present disclosure may be considerednon-irritating and non-inflammatory if it does not cause more than a 25%release of markers of cell damage or expression of pro-inflammatorymediators above that caused by a negative control (e.g., syntheticTLR2/6 ligand). In some embodiments, the compositions of the presentdisclosure may be considered non-irritating and non-inflammatory togenital skin and/or mucosa if application of the composition to thespecific region does not increase inflammasome and/or cytokineproduction (e.g., by more than about 1%, more than about 5%, more thanabout 10%, more than about 15%, more than about 20%, more than about25%) as compared to control. For example, a control reference level maybe the level of the indicated biomarker expressed as an average of thelevel of the biomarker from samples taken from a control population ofhealthy subjects. In some embodiments, a control reference level may bethe level of the indicated biomarker expressed as an average of thelevel of the biomarker measured from a subject given a controlformulation. Suitable samples or references for determining referencelevels include healthy cells. In some embodiments, the reference todetermine the reference level of an indicated biomarker may be a derivedfrom the subject, a healthy subject, or a population of subjects.

Healthy mucin and mucus, and mucin-regulating enzyme production(glycocidases), from the vaginal epithelial cells can be determinedfollowing exposure of genital tissue or fluid to compositions used forcleaning and lubricating the anogenital region. Current urogenitaland/or anogenital products can damage the natural mucus protectionbarrier of the surface of genital skin or mucosa of the vagina, penisand urethra, through altering mucin production and enhancingmucin-degradation. In particular, mucin degradation can occur followingexposure to certain pathogenic bacteria or to certain ingredients (e.g.carbomer and oils) commonly found in products used in the genitalregion. Mucin quality from the vagina can be determined followingexposure to a composition of the present disclosure by testing CVFsamples collected using a SoftCup or similar menstrual cup devicecovering the base of the cervix and performing ELISA assay (enzymelinked immunosorbent assay) to measure mucins and ELLA assay (enzymelinked lectin assay) to measure carbohydrate structures as described inMoncla et al. (PLOS One 11:e0158687, 2016, incorporated herein byreference in its entirety). Glycosidase assays can be performed tomeasure enzyme specific activity as described in Moncla et al. (PLOS One11:e0158687, 2016, incorporated herein by reference in its entirety). Acomposition of the present disclosure is considered non-harmful togenital mucin and mucus if it does not inhibit mucus viscosity orproduction (e.g., by more than about 1%, about 5%, about 10%, about 15%,about 20%, about 25%, about 30%, or about 35%), or increase productionof glycocidases (e.g., by more than about 1%, about 5%, about 10%, about15%, about 20%, about 25%, about 30%, or about 35%) (see, Moncla et al.,PLOS One 11:e0158687, 2016, incorporated herein by reference in itsentirety). Compositions of the present disclosure may also be tested fortheir effects in vaginal infection susceptibility models, such as amouse genital herpes transmission model (see, e.g., Moench et al., BMCInfect. Dis. 10:331, 2010). Increased susceptibility to infectiontransmission may be caused by damage to vaginal epithelial cells.

Effects of topical compositions on tissue viability using tissue models(e.g. human explants or cell monolayers) may also be assessed using theMTT colorimetric assay technique. The MTT assay is a colorimetric assayfor assessing cell metabolic activity. NAD(P)H-dependent cellularoxidoreductase enzymes may, under defined conditions, reflect the numberof viable cells present. These enzymes are capable of reducing thetetrazolium dye MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide to its insoluble formazan, which has a purple color. The MTTassay may be used to measure a composition's cytotoxicity or effect oncell viability (Ayehunie et al., 2011).

In addition, oxidative stress and antioxidant potential of the tissuescan be determined by common methods, such as a TBARS assay to evaluatethe impact of various embodiments on tissue health. Because reactiveoxygen species (ROS) have extremely short half-lives, they are difficultto measure directly. Instead, several products of the damage produced byoxidative stress, such as thiobarbituric acid reactive substances(TBARS), can be measured. TBARS are formed as a byproduct of lipidperoxidation (i.e., as degradation products of fat), which can bedetected by the TBARS assay using thiobarbituric acid as a reagent.

The in vivo rabbit vaginal irritation (RVI) model may also be used toassess the irritation and inflammatory characteristics of a formulation(Eckstein et al., J. Reprod. Fertil. 20:85, 1969). This model is basedon macroscopic observations of erythema, edema and ulceration andhistopathologic analysis of the tissues collected after exposure of theanimals to the test materials. A non-irritating and safe composition ofthis disclosure would show no negative macroscopic or histopathologiceffects as compared to a control vehicle. An expanded RVI model having aquantitative nuclease protection assay (qNPA) to quantify mRNA levels of25 genes representing leukocyte differentiation markers, toll-likereceptors (TLR), cytokines, chemokines, epithelial repair, microbicidaland vascular markers has also been described (see, Fichorova et al.,Toxicol. Appl. Pharmacol. 285:198, 2015).

Sensitization tests may be used evaluate the potential of a compositionof the present disclosure to cause a sensitizing effect or allergenicreaction in a subject over an extended period of exposure. Exemplarytests include Guinea pig tests, such as the Magnusson-Kligman guinea pigmaximization test (J. Invest. Dermatol. 52:268, 1969), the occludedpatch test of Buehler (Arch. Dermatol. 91:171, 1965), and the openepicutaneous test (Kero et al., Contact Dermatitis 6:341, 1980). Amurine local lymph node assay (LLNA) is another method for theidentification of skin sensitizing chemicals. In contrast to guinea pigtests, this assay relies on measurement of events induced during theinduction phase of skin sensitization, specifically lymphocyteproliferation in the draining lymph nodes which is a hallmark of a skinsensitization response, rather than the elicitation phase (Kimber etal., Contact Dermatitis 21:215, 1989; Basketter et al., Food Chem.Toxicol. 34: 985, 1996). The human repeat-insult patch test (HRIPT) maybe performed as a confirmatory test in the safety evaluation of skinsensitizers. Sensitization is a process by which humans developincreased allergic responses to a substance over time through repeatedexposure to that substance. It is different from irritation because itinvolves an immune response. Skin sensitization reactions are usuallycharacterized by erythema coupled with one or more of various dermalsequelae, such as edema, papules, vesicles, bullae, and/or pruritus(McNamee et al., Regul. Toxicol. Pharmacol. 52:24, 2008).

In yet another example, in vivo colposcopy exams of women following useof compositions of the present disclosure can identify signs ofinflammation or irritation. User surveys can also be used for scoring ofsymptoms of the same (Van Damme et al., Lancet 360:971, 2002; Bunge etal., J. Acquir. Immune Defic. Syndr. 60:337, 2012).

Irritating topical products may trigger the release of pro-inflammatorycytokines (e.g., TLR, IL-1, IL-6, IL-8, TNF-α, IFN-γ, IL-17) andinflammasomes (e.g., NLRP3 and NLRC4). Cytokines and inflammasomes canbe measured using an enzyme-linked immunosorbent assay (ELISA),quantitative PCR, or other molecular assay. A product is considerednon-inflammatory if it does not cause increased expression ofpro-inflammatory cytokines or inflammasomes (Rabeony et al., Eur. J.Immunol. 45:2847, 2015).

In certain embodiments, the compositions of the present disclosure donot induce irritation or inflammation potential in the anogenital regionsubject greater than about 0.5%, about 1%, about 2%, about 3%, about 4%,about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%,about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%,about 25%, about 26%, about 27%, about 28%, about 29%, or about 30% ascompared to an untreated control subject, preferably as measured usingthe slug mucosal irritation test of Adriaens et al. (2008).

In certain embodiments, the compositions of the present disclosure donot impact sperm viability or function. In certain embodiments,compositions of the present disclosure are designed to mimic the vaginalenvironment during a woman's fertile window around ovulation and notnegatively impact sperm viability or function. Assays or models forassessing sperm survival and function, include for example, spermmotility assays (e.g., subjective or computer assisted), sperm viabilitystudies, in vitro fertilization and embryo development animal models,membrane integrity of sperm, survival time in culture, cervical mucuspenetration, lipid peroxidation, capacitation, zona recognition,acrosome reaction and sperm-oocyte fusion, and sperm chromatin testing(reviewed in, e.g., Vasan, Indian J Urol. 27:41, 2011; Oehninger et al.,Fertil. Steril. 102:1528, 2014; Mortimer et al., Hum. Reprod. Update 19(Suppl 1):i1-i45; 2013, each of which is incorporated by reference inits entirety). Additional testing can include post-coital testing toevaluate sperm presence in the cervical canal, and even pregnancyoutcomes in an animal model or among women in a clinical trial.

Sperm motility is one function that may be used to assess sperm functionand thus fertilization potential. Motility of sperm is expressed as thetotal percent of motile sperm, the total percent of progressively motilesperm (swimming forward), or the speed of sperm that are progressivelymotile. These measurements may be made by a variety of assays, but areconveniently assayed in one of two ways. Either a subjective visualdetermination is made using a phase contrast microscope when the spermare placed in a hemocytometer or on a microscope slide, or a computerassisted semen analyzer is used. Under phase contrast microscopy, motileand total sperm counts are made and speed is assessed as fast, medium orslow. A more specific of sperm motility is motility grade, where themotility of sperm is divided into four different grades (Cooper et al.,Human Reprod. Update 16:231, 2010, incorporated by reference in itsentirety). Grade A refers to sperm with progressive motility that arethe strongest and swim fastest in a straight line. Grade B refers tosperm with non-linear motility; that move forward but tend to travel ina more curved or crooked motion. Grade C sperm have non-progressivemotility in that they do not move forward despite tail movement. Grade Dsperm are immotile. Using a computer assisted semen analyzer (such asIVOS Hamilton Thorne, Beverly, Mass.), the motility characteristics ofindividual sperm cells in a sample are objectively determined (Hum.Reprod. 13:142, 1998). Briefly, a sperm sample is placed onto a slide orchamber designed for the analyzer. The analyzer tracks individual spermcells and determines motility and velocity of the sperm. Data may beexpressed as percent of total motility, and measurements are obtainedfor path velocity and track speed as well. It is known that the velocityof sperm is often impacted by the viscosity of a medium and separatefrom the toxicology of that medium as described in J Elgeti, et al.,Biophys J 99 (2010): 1018-1026, hereby incorporated by reference in itsentirety.

The Human sperm survival assay is typically used in human in vitrofertilization (IVF) programs as described in C De Jonge et al., J Androl24 (2003): 16-18, and A Hossain, et al., Adv Urol 2010 (2010): 136898,each of which is hereby incorporated by reference in its entirety. Ithas also been proposed as a sensitive cytotoxic assay for any topicalproducts, as sperm show damage before other cell monolayer types. Thetest requires sperm suspension culture with 10% product for 24 hours andevaluation of the percent motile, progressive motility and/or totalmotility of sperm at the end of culture in climate controlled settings.The test can also be replicated with bull sperm for controlling forindividual sperm sample effect. In certain embodiments, the compositionsof the present disclosure are considered not toxic to sperm ifapplication to a subject does not cause a decrease in sperm survivalgreater than about 0.5%, greater than about 1%, greater than about 2%,greater than about 3%, greater than about 4%, greater than about 5%,greater than about 6%, greater than about 7%, greater than about 8%,greater than about 9%, greater than about 10%, greater than about 11%,greater than about 12%, greater than about 13%, greater than about 14%,greater than about 15%, or greater than 20% as compared to control.

Sperm viability can be measured using several different methods. By wayof example, two of these methods are staining with membrane exclusionstains and measurement of ATP levels. Briefly, a sample of sperm isincubated with a viable dye, such as Hoechst 33258 or eosin-nigrosinstain. Cells are placed in a hemocytometer and examined microscopically.Dead sperm with disrupted membranes stain with these dyes. The number ofcells that are unstained is divided by the total number of cells countedto give the percent live cells. ATP levels in a sperm sample aremeasured by lysing the sperm and incubating the lysate with luciferase,an enzyme obtained from fireflies, which fluoresces in the presence ofATP. The fluorescence is measured in a luminometer. The amount offluorescence in the sample is compared to the amount of fluorescence ina standard curve allowing a determination of the number of live spermpresent in the sample.

Membrane integrity of sperm is typically assayed by a hypo-osmotic swelltest which measures the ability of sperm to pump water or salts ifexposed to non-isotonic environments. Briefly, in the hypo-osmotic swelltest, sperm are suspended in a solution of 75 mM fructose and 25 mMsodium citrate, which is a hypo-osmotic (150 mOsm) solution. Sperm withintact, healthy membranes pump salt out of the cell causing themembranes to shrink as the cell grows smaller. The sperm tail curlsinside this tighter membrane. Thus, sperm with curled tail are countedas live, healthy sperm with normal membranes. When compared to the totalnumber of sperm present, a percent of functional sperm may beestablished.

The degree of membrane integrity is preferably determined by lipidperoxidation (LPO) measurements, which assess sperm membrane damagegenerated by free radicals released during handling. Lipid membraneperoxidation is assayed by incubating sperm with ferrous sulfate andascorbic acid for one hour in a 37° C. water bath. Proteins areprecipitated with ice-cold trichloroacetic acid. The supernatant iscollected by centrifugation and reacted by boiling with thiobarbituricacid and NaOH. The resultant malondialdehyde (MDA) formation isquantified by measuring absorbance at 534 nm as compared to an MDAstandard (Bell et al., J. Andrology 14:472, 1993, incorporated herein byreference in its entirety). LPO is expressed as nM MDA/10⁸ sperm. Acomposition of the present disclosure has a stabilizing effect on spermif exposure results in decreased LPO production.

The stability of chromatin DNA is assayed using the sperm chromatinstructure assay (SCSA). Sperm cells are very sensitive to oxidativestress resulting in sperm chromatin (DNA) damage (Whittington et al.,Int. J. Andrology 22:236, 1999; Pasqualotto et al., Fertility andSterility 73:459, 2000; Kodama et al., Fertility and Sterility 68:519,1997, each of which is incorporated herein by reference in itsentirety). This damage can be profound in sperm cells because theycontain little to no mechanisms to repair DNA damage after it occurs.The sperm chromatin assay is based on the metachromatic staining ofsingle and double stranded DNA by acridine orange stain (Evenson et al.,Human Reprod. 14:1039, 1999; Evenson et al., J. Andrology 23:25, 2002,each of which is incorporated by reference in its entirety). Excitationwith an argon laser causes acridine orange intercalated indouble-stranded DNA to emit a green fluorescence, whereas redfluorescence is emitted by single strand DNA. The extent of DNAdenaturation in a sample is expressed as a and calculated by the formulaα=red/(red+green). The endpoint measurement is DNA Fragmentation Index(DFI). A DFI of <15% DFI indicates excellent to good sperm DNAintegrity. Fresh sperm samples are incubated for a period of time in thepresence of a test composition, flash frozen, and subsequently assayedfor DNA breakage (see, e.g., Evenson et al., J. Androl. 23:25, 2002,incorporated herein by reference in its entirety). Other DNA assays forthe stability of chromatin DNA include the terminal deoxynucleotidyltransferase-mediated fluorescein-dUTP nick end labelling (TUNEL) test;COMET assay and Sperm Chromatin Dispersion as disclosed in D Evenson,Anim Reprod Sci 169 (2016): 56-75, hereby incorporated by reference inits entirety.

In vitro fertilization rates are determined by measuring the percentfertilization of oocytes in vitro in an animal model such as bovine ormurine model. For example, maturing bovine oocytes are cultured in vitroin M199 medium plus 7.5% fetal calf serum and 50 μg/ml luteinizinghormone for 22 hours. Following culture for 4 hours, the sperm arechemically capacitated by adding 10 IU of heparin and incubated withbovine oocytes for 24 hours. At the end of the incubation, oocytes arestained with an aceto-orcein stain or equivalent to determine thepercent oocytes fertilized. Alternatively, fertilized oocytes may beleft in culture for 2 days, during which division occurs and the numberof cleaving embryos (i.e., 2 or more cells) is counted.

Survival time in culture of sperm (time to loss of motility) is anotherconvenient method of establishing sperm function. Briefly, an aliquot ofsperm is placed in culture medium, such as Tyrode's medium, pH 7.4 andincubated at 37° C., 5% CO₂, in a humidified atmosphere. At timedintervals, for example every 2 hours, the percentage of motile sperm inthe culture is determined by visual analysis using an invertedmicroscope or with a computer assisted sperm analyzer. As an endpoint, asperm sample is considered no longer viable when less than 5% of thecells have progressive motility.

Another parameter of sperm function is the ability to of sperm to swimup into a column of cervical mucus or substitute (reviewed in Ola etal., Hum. Reprod. 18:1037-1046, 2003, incorporated by reference in itsentirety). This cervical mucus penetration test can be done either invitro or in vivo. Sperm are placed at one end of the track and thedistance that sperm have penetrated into the mucus after a given timeperiod is determined. Cervical mucus penetration studies offer valuablebiocompatibility data for devices that are used for reproductivepurposes. The bovine cervical mucus penetration study is an excellent invitro assay to evaluate sperm penetration into cervical mucus. Bergmanet al. (Fertility and Sterility 36:363-367, 1981, incorporated herein byreference in its entirety) found excellent correlation between spermpenetration into frozen bovine cervical mucus and fresh human cervicalmucus (r=0.98) due to the similarity of human and bovine cervical mucusrheological and biophysical make-up (Bergman et al., Fertil. Steril.36:363-367, 1981; Keel et al., Arch. Androl. 44:109-115, 2000, each ofwhich is incorporated herein by reference in its entirety). These assaysmay be used to evaluate cytotoxicity of aspects of the presentdisclosure, such as applicators, compositions, and kits, by incubatingthe embodiment (e.g. applicator) in a container with a mixture of thecomposition (e.g. 10% composition by volume of the solution), cervicalmucus and a sperm solution, in order to determine toxicity of leachedproducts from the device on sperm penetration into cervical mucus.Toxicity of sperm penetration can also be measured by placing thecompositions of the present disclosure in an applicator (e.g., anapplicator as described herein), and incubating the composition with theapplicator for a set time (e.g. more than 10 min, more than 20 min, morethan 25 min, 30 min), whereafter the incubated composition is mixed withcervical mucus and sperm to form a solution. This solution may be usedto evaluate subsequent sperm penetration into mucus, to determineeffects of sperm exposed to the composition and/or identify the presenceof any leached chemicals from the applicator into the composition. Incertain embodiments, the compositions of the present disclosure are nottoxic to sperm if there is not a decrease in sperm penetration ofgreater than about 0.5%, of greater than about 1%, of greater than about2%, of greater than about 3%, of greater than about 4%, of greater thanabout 5%, of greater than about 6%, of greater than about 7%, of greaterthan about 8%, of greater than about 9%, of greater than about 10%, ofgreater than about 11%, of greater than about 12%, of greater than about13%, of greater than about 14%, of greater than about 15% of control.

Alternatively, sperm penetration of mucus may be measured in vivo inwomen. At various times post-coitus, a sample of cervical mucus isremoved and examined microscopically for the number of sperm present inthe sample. In the post-coital test, improved sperm function isestablished if more sperm with faster velocity are seen in the mucussample after exposure to a composition of the present disclosure versusa sample of mucus from the patient after exposure to a controllubricant.

Other assays of sperm function potential include the sperm penetrationand hemizona assays. In the sperm penetration assay, the ability ofsperm to penetrate into an oocyte is measured. Briefly, commerciallyavailable zona free hamster oocytes are used (EmbryoTech Laboratories,Haverhill, Mass.). Hamster oocytes are suitable in this assay for spermof any species. Capacitated sperm, such as those cultured with bovineserum albumin for 18 hours, are incubated for 3 hours with the hamsteroocytes. Following incubation, oocytes are stained with acetolacmoid orequivalent stain and the number of sperm penetrating each oocyte iscounted microscopically. A hemizona assay measures the ability of spermto undergo capacitation and bind to an oocyte. Briefly, in this assay,live normal sperm are incubated in media with bovine serum albumin,which triggers capacitation. Sperm are then incubated with dead oocyteswhich are surrounded by the zona pellucida, an acellular coating ofoocytes. Capacitated sperm bind to the zona and the number of spermbinding is counted microscopically.

In certain embodiments, a composition of the present disclosure isnon-toxic to sperm if following exposure to a 10% solution of thecomposition, sperm retain at least 80%, at least 85%, at least 90%, orat least 95% motility as compared to sperm exposed to a control medium.

In certain embodiments, the topical, isotonic composition of the presentdisclosure: (i) is non-irritating to the urogenital and/or anogenitalmucosa or skin; (ii) does not promote growth of pathogenic bacteria ofthe urogenital and/or anogenital region; (iii) does not reduce thehealthy microbiota of the genital region more than about 25%; (iv) doesnot disrupt or reduce mucin production by the urogenital and/oranogenital mucosa or skin; (v) does not cause more than about a 20%increase in inflammation of the urogenital and/or anogenital mucosaand/or the skin genital tissue; (vi) does not disrupt genital fluidfunction; (vii) is non-toxic to sperm; (viv) has a prebiotic effect onLactobacillus species growth found in the genital tissues; (viv)decreases vaginal, bacterial, fungal, or viral infection rates by 5% ormore; or any combination thereof

Methods of Use

Disruption of the genital microbiota can lead to problems with genitaldiscomfort, odor, burning, dyspareunia, scarring, irritation,inflammation, infection, infertility, inferior reproductive outcomes,and increase risk of autoimmune disease, sexually transmittedinfections, cancer, and/or dementia. Disruption of the genitalmicrobiota can be caused by a variety of factors including diseases(e.g., diabetes); lichen sclerosis; autoimmune diseases such aspsoriasis; interstitial cystitis; hormonal changes (e.g., pregnancy,puberty, post-partum, menopause, andropause); urinary incontinence;fertility treatments; gender reassignment therapy (e.g., hormonereplacement therapy and/or sex reassignment surgery); sexual initiationand introduction of new sexual partners; childbirth; immunosuppression;cancer therapy; use of alkaline soaps and washes; or use of irritatinglubricants, douches, creams or medications. Topical compositions of thepresent disclosure are pH and isotonic specific to the urogenital and/oranogenital regions and/or genital fluids; support, promote or enhancespecific genital microbiota; are non-irritating to the urogenital and/oranogenital region; do not disrupt normal anogenital mucin production;and thus, are useful for a variety of applications in the urogenitaland/or anogenital region.

Without, wishing to be bound by theory, the compositions of the presentdisclosure improve Lactobacillus dominance in the genital tissues inwomen of reproductive age and post-menopausal age; and girls prior topuberty. Beneficial genital microbiomes also decrease pathogens, lowervaginal pH, and/or improve Female Sexual Function Index scores withregular use. These compositions may decrease vaginal infection ratesand/or urinary tract infections, especially in women in settings proneto exacerbation of these symptoms. Patients in need of thesecompositions may be women including women in low resource settings suchas military deployment, natural disaster setting, or low-incomecommunities, older women with dementia, pre-pubescent girls with lichensclerosus, women with interstitial cystitis, and women with provokedvestibulodynia.

In most embodiments, compositions are provided which are pH balanced fordifferent stages in a woman's cycle or life. For example, thecompositions may have an acidic pH of (e.g., from 4-5, about 4.5, about4.8) for most stages of a woman's cycle or life. In other embodiments,the compositions may have a closer to neutral or alkaline pH (e.g., fromabout 6 to about 7 from about 6.5 to about 6.8, from about 6.5 to about7, from about 6.7 to about 6.9, about 6.8) for use during ovulation forfertility enhancement.

In one aspect, the present disclosure provides a method of hydrating ormoisturizing the urogenital and/or anogenital region of a subjectcomprising topically administering an effective amount of a topical,isotonic composition of the present disclosure to the urogenital and/oranogenital region of the subject. The method may be used to hydrate ormoisturize the genital tissues, including the perineum, penis orvulva/vagina. Irritation of the skin of the penis and disruption ofhealthy mucin secretions that moisten and protect the penis can increasedryness, roughness and inflammation resulting in pain and discomfort.Vaginal intercourse, which introduces the penile surface to a very lowpH environment can irritate the skin of the penis. Similarly, irritationof the vagina skin around the vulva can occur following ejaculation ofhigh pH and hypertonic semen into/onto the female genital region,leading to burning and post-coital pain. Continuous washing and cleaningof the urogenital and/or anogenital region, particularly with alkalinesoaps or washes, can also irritate and dry out the urogenital and/oranogenital region. By hydrating or moisturizing the urogenital and/oranogenital region and buffering pH the topical, isotonic compositions ofthe present disclosure may be used to promote, enhance, protect theanogenital epithelium and microbiota. In certain embodiments, topical,isotonic compositions further comprise an additional therapeutic agent,such as a topical pain-relieving agent.

In another aspect, the present disclosure provides a method ofconditioning the skin of the urogenital and/or anogenital region of asubject comprising topically administering an effective amount of atopical, isotonic composition of the present disclosure to theurogenital and/or anogenital region of the subject. The decrease-insystemic estrogen levels that occurs during menopause and pregnancy cancause dry, thin, friable vulvar skin and vaginal dryness. As men age,senescence of penile epithelial tissues occurs similar to elsewhere inthe body. However, high content of elastic fibers and collagen in thepenis is required to allow for the repeated and regular expansion andretraction in size of the penis. The topical, isotonic compositions ofthe present disclosure may be used for genital skin and mucosaconditioning and genital microbiome support, particularly in an agingsubject (e.g., about 50 years or greater, about 60 years or greater,about 70 years or greater, about 80 years or greater, about 90 years orgreater, about 100 years or greater) or a subject experiencing menopauseor andropause. In certain embodiments, the topical, isotonic compositionfurther comprises an additional therapeutic agent, such as a hormone,erectile dysfunction treatment or erectile enhancement drug, prematureejaculation drug, or a combination thereof. Furthermore, diseases suchas lichen sclerosus, diabetes and auto-immune diseases can make genitalepithelial surfaces highly reactive to inflammation and irritation. Thetopical, isotonic compositions of the present disclosure may be used forgenital skin and mucosa conditioning and genital microbiome support in asubject with systemic or localized epithelial diseases.

In another aspect, the present disclosure provides a method oflubricating the urogenital and/or anogenital region of a subjectcomprising topically administering an effective amount of a topical,isotonic composition of the present disclosure to the urogenital and/orurogenital and/or anogenital region of the subject. In certainembodiments, the urogenital and/or anogenital region is the perineum,vagina, vulva, clitoris, penis, scrotum, or anus. The topical, isotoniccomposition may be administered to the urogenital and/or anogenitalregion of the subject prior to, during, and/or after sexual activity.Sexual activity includes oral sex, penetrative sex (e.g., vaginalintercourse, anal intercourse), non-penetrative sex, genital contactwith a body part (e.g., hand, foot), genital contact with an object(e.g., sex toy), masturbation, dry humping (genital rubbing), or anycombination thereof. In certain embodiments, the topical, isotoniccomposition is administered to the urogenital and/or anogenital regionof the subject for use with a sex toy. In additional embodiments, thetopical, isotonic composition is administered to a medical device,contraceptive device, or sex toy in alternative or in addition toadministration to the urogenital and/or anogenital region. For example,the topical, isotonic composition may be administered to the interior orexterior of a condom, to the interior or exterior of a sex toy, to theexterior of a menstrual cup, to the exterior of a diaphragm, or to theexterior of a vaginal ultrasound or speculum prior to contact with thesubject's urogenital and/or anogenital region. During vaginal,heterosexual intercourse, the penis is introduced to low pH vaginalsecretions. Sexual intercourse frequently leads to dysbiosis andproduction of odorous amines from anaerobic bacteria. Most commerciallubricants contain irritating and damaging ingredients, such asphoto-oxidized oils, silicones, and chemicals, and are hypertonic (e.g.,five times the level of physiologic fluids), which can further irritatethe urogenital and/or anogenital region when used during sexualactivity, such as intercourse or masturbation. The topical, isotoniccompositions of the present disclosure may provide urogenital and/oranogenital lubrication and genital microbiome support without irritatingthe urogenital and/or anogenital region in a subject. In certainembodiments, the topical, isotonic composition further comprises anadditional therapeutic agent, such as a hormone, a drug for treatingvaginal atrophy (e.g., genitourinary syndrome of menopause), a drug forenhancing sexual responsiveness (e.g. orgasmic latency), erectiledysfunction treatment or erectile enhancement drug, a drug for prematureejaculation, an agent that enhances vasodilation, a microbicide toprevent STDs, a chemical contraceptive, or a combination thereof.

Thus, in another aspect, the present disclosure provides a method ofcleaning the urogenital and/or anogenital region of a subject comprisingtopically administering an effective amount of a topical, isotoniccomposition of the present disclosure to the urogenital and/oranogenital region of the subject. The use of harsh alkaline soaps andwashes (pH˜8-11) to clean the urogenital and/or anogenital region (e.g.,penis or vulva/vagina) can disrupt the microbiota, leading to unpleasantgenital odor, irritation, and BV. Semen, sweat, and some lubricants alsohave an alkaline pH. Many commercial diaper wipes have very acidic pH(e.g., pH 2.8) or alkaline pH (e.g., pH 8), which can depress orelevate, respectively, the normal skin pH of the urogenital and/oranogenital area following use (Priestly et al., Pediatr. Dermatol.13:14, 1996). The normal penile skin has a pH of about 4.5-6. The normalpH of the vulva/vagina is generally acidic due to the Lactobacilli inthe microbiota, although vulvar pH is higher than that of vagina.However, the pH of the female genital region varies depending on thelife stage and menstrual cycle stage. During most of the menstrualcycle, the female genital region is often at a pH of about 4.5. However,during ovulation, the cervical secretions becomes more neutral at aboutpH 6 to about pH 7 (which facilitates rapid sperm transport through thecervix). During pregnancy and menopause, the pH of these secretions isoften at about 5. These states are prone to vaginal dysbiosis.Postmenopausal women have high rates of BV (approaching 50%) and BVrates increase over time after menopause. Exposure to alkaline pHassociated with BV or dysbiosis can disrupt the genital microbiota,leading to increased pathogenic bacteria growth and production ofoffensive biogenic amines, increased vaginal symptoms, sexuallytransmitted diseases, and cancers. Individuals who have increasedsweating in the groin area due to inherent physiology, weight gain orfrequent exercise, in combination with an imbalanced penile or vaginalmicrobiome, can also have increased offensive genital odor that canimpact quality of life. Moreover, genital odor complaints and dysbiosisare more significant in uncircumcised men due to the collection ofsecretions in the preputial fornix. The topical compositions of thepresent disclosure are non-irritating, and pH and isotonic specific tothe urogenital and/or anogenital region of interest.

In another aspect, the present disclosure provides a method ofdecreasing irritation or inflammation of the urogenital and/oranogenital region of a subject comprising topically administering aneffective amount of a topical, isotonic composition of the presentdisclosure to the urogenital and/or anogenital region of the subject.

In another aspect, the present disclosure provides a method ofpreventing cervical cancer, wherein the compositions of the presentdisclosure are administered to a subject in need thereof.Administrations of these compositions may serve as a prebiotic forbeneficial genital microbiota growth, and enhancing high risk HPVclearance.

In another aspect, the present disclosure provides a method ofdecreasing bothersome vaginal symptoms in older women withpostmenopausal BV, urinary incontinence, pelvic floor prolapse or otherforms of pelvic floor disease wherein the compositions of the presentdisclosure are administered to a subject in need thereof, includingoptional concurrent use with a vaginal pessary for prolapse management.

In another aspect, the present disclosure provides a method ofdecreasing bothersome penile skin conditions, resulting in redness,roughness, scabbing, scaling, itching, and odor wherein the compositionsof the present disclosure are administered to a subject in need thereof.

In another aspect, the present disclosure provides a method of enhancingsexual enjoyment and pleasure, by decreasing friction, irritation, painand orgasm latency wherein the compositions of the present disclosureare administered to a subject in need thereof.

In another aspect, the present disclosure provides a method ofdecreasing Type 1 diabetes in offspring by enhanced Lactobacillusdominance in the mother's vagina prior to birth wherein the compositionsof the present disclosure are administered to the mother in needthereof. The connection between diseases such as Type 1 diabetes inoffspring have been associated with the genital microbiome of the motheras disclosed in M. Tejesvi, et al., Sci Rep 9 (2019): 959, herebyincorporated by reference in its entirety and particularly in relationto the connection between the vaginal microbiome and Type 1 diabetes inmembers of a dyad.

In another aspect, the present disclosure provides a method ofoptimizing, establishing, and maintaining a healthy microbiome followinggender reassignment surgery comprising administration of a compositionof the present disclosure to a subject in need thereof.

In yet another aspect, the present disclosure provides a method ofsupporting, enhancing, or promoting the genital microbiota of a subjectcomprising topically administering an effective amount of a topical,isotonic composition of the present disclosure to the genital region ofthe subject. In certain embodiments, the topical, isotonic compositionis prebiotic for Lactobacillus species growth. In certain embodiments,the topical, isotonic composition may further comprise at least oneprobiotic bacterial species (e.g., Lactobacillus species).

In yet another aspect, the present disclosure provides a system for amethod of hydrating, conditioning, cleansing or lubricating theurogenital and/or anogenital region of one member or both members of asexual dyad throughout a reproductive cycle, in order to enhancereproductive outcomes. In certain embodiments, the sexual dyad comprisesat least one female. In certain embodiments, the sexual dyad is aheterosexual dyad. The reproductive cycle refers to the menstrual cycleor hormone changes required for the production of an oocyte (ovariancycle) and preparation of the uterus for pregnancy (uterine cycle). Theovarian cycle includes the follicular phase (pre-ovulatory phase),ovulation, and luteal phase (post-ovulatory phase). The uterine cycleincludes menses, proliferative phase, and secretory phase. Embodimentsof this system provide urogenital and/or anogenital cleansing andlubrication at a pH and tonicity consistent with the healthy function ofthe penile, clitoral, vaginal, and/or vulvar ecosystem(s), theurogenital ecosystems, cervical fluids, sperm, and semen. The system mayprovide urogenital and/or anogenital cleansing and lubrication at a pHand osmolality consistent with the healthy function of cervical fluids,and sperm in the vagina post-ejaculation. Specific embodiments ofcompositions of the present disclosure are used by one or both membersof the sexual dyad during the non-fertile portion of the cycle (e.g.,the follicular phase) (Step 1). In certain embodiments, the follicularphase refers to the period of time from day 0 of menstrual cycle up tothe fertile window. These compositions are used for cleansing, and as aleave-in conditioner to optimize the healthy genital microbiome, and asa coital lubricant applied to the vaginal canal prior to intercourse orsexual activity. The compositions are buffered, muco-adhesive and have atonicity of about 150 mOsmo/kg and a pH of about 4.5. Other embodimentsof compositions of the present disclosure are then used during thefertile phase (Step 2). As used herein the fertile window or fertilephase includes the luteinizing hormone surge that occurs about 24 toabout 36 hours before ovulation and ovulation. In certain embodiments,the fertile phase refers to the period of about 6 days, about 5 days,about 4 days, about 3 days, about 2 days, or about 1 day beforeovulation through about 24 hours or about 36 hours after ovulation.These compositions are used for cleansing, and as a leave-in conditionerto enhance the healthy genital microbiome, and as a coital lubricantapplied to the vaginal canal prior to intercourse or sexual activity.These compositions have about a tonicity of about 240 mOsmo/kg and a pHof from about 5 to about 6.8. In certain embodiments, sperm function isprotected at the lower pH of the vagina by compositions with enhancedanti-oxidant activity, and improved cervical mucus quality to supportrapid transport of sperm out of the acidic vaginal environment.Protecting post-ejaculatory sperm during transport through a compositionwhich is at a lower pH (e.g. pH=5) than that of typical cervical mucus(e.g. pH 6.8), may limit subsequent dysbiosis and BV risk in the mother.In certain embodiments, from about 36 hours following ovulation, thecompositions from Step 2 are discontinued (e.g., luteal phase) andcompositions from Step 1 are used to facilitate return of the vaginalcanal to an acidic, hypotonic environment. In certain embodiments, theperiod of time outside the fertile window, including before, after, orboth before and after the fertile window is referred to as thenon-fertile phase or time. Some current fertility lubricants weredesigned to protect sperm during transport through the vagina and cervixto the egg. However, the higher pH (usually >7) and higher tonicity(three times the normal CVF) of these products could disrupt healthyvaginal cleaning and function, as they are used in amounts equal to thatof the ejaculate, thereby disrupting the normal balance of CVF-sementhat occurs during intercourse. Normally, the admix of CVF and semenresults in a pH and tonicity almost perfectly matched to the pH andtonicity of blood and body tissues. Compositions and systems of thepresent disclosure do not disrupt this elegant balance, which mayprotect the female from dysbiosis, e.g., during any resulting pregnancy.In certain embodiments, the compositions of the present disclosureadjust the postcoital admix of fluids (e.g., CVF, semen) in the vaginato a pH of about 6 and tonicity of about 240 mOsmo/kg. In certainembodiments, the compositions of the present disclosure adjust thepostcoital admix of fluids (e.g., CVF, semen) in the vagina to a pH offrom 4 to about 6 or from 4.5 to about 5.5 or about 5. These adjustmentsmay adjust the tonicity of the environment to about 240 mOsmo/kg.Typically, perm function is preserved during exposure to the lower pH bythe composition for the 30 minutes or required time for sperm transportthrough the human vagina. Such mechanisms are discussed in F. Nakano,Medical Express 2 (2015): ISSN 2318-8111, hereby incorporated byreference in its entirety.

In certain embodiments, the pH of the topical, isotonic composition isformulated at a pH to match to the normal, physiological genital fluidpH (e.g., CVF, urethral secretions, semen) or at a pH appropriate forthe particular method of use. In certain embodiments, the pH of thetopical, isotonic composition ranges from about 3.5 to about 6.8. In aparticular embodiment, the pH of the topical, isotonic composition isabout 4.5, 5, about 5.5, about 6.5, or about 6.8. Topical, isotoniccompositions having a pH ranging from about 3.5 to about 6.8, areparticularly suited for administration to a subject that is aged aboutfrom puberty to menopause/andropause, e.g., about 18 years of age toabout 50 years of age, about 18 years of age to about 55 years of age,about 18 years of age to about 60 years of age, or a child at least oneyear old. In certain embodiments, the pH of the topical, isotoniccomposition ranges from about 5 to about 8. In a particular embodiment,the pH of the topical, isotonic composition is about 6 or about 6.8.Topical, isotonic compositions having a pH ranging from about 5 to about7 are particularly suited for administration to an infant aged from 0 toabout 12 months old, a senior adult of at least about 60 years of age,an adult (male or female) of reproductive age (e.g., ranging from about18 years to about 50 years) who is actively trying to conceive, or bothmembers of a heterosexual dyad which is actively trying to conceive.

The methods provided in the present disclosure may be used on amammalian subject (e.g., bovine, canine, feline, equine, porcine, ovine,avian, rodent, lagomorph, caprine, non-human primate), preferably ahuman subject. In certain embodiments, the subject is a male, a female,an intersex subject, a non-binary gendered subject, or a subject of anyother gender designation. In certain embodiments, the subject is aninfant, a child, or an adult. In certain embodiments, the subject is anadult male of reproductive age (e.g., ranging about 18 years to about 50years) that is trying to conceive or adult female of reproductive age(e.g., ranging from about 18 years to about 50 years) that is trying toconceive. In certain embodiments, the subject is a female in menopauseor male in andropause. In certain embodiments, the subject is an infant(aged from 0 to about 12 months old), a child at least 1 year old; anadult ranging from about 18 years to about 50 years of age, about 18 toabout 55 years of age, or about 18 to about 60 years of age; or a senioradult of at least about 50 years, at least about 55 years, at leastabout 60 years, or at least about 65 years of age. In certainembodiments, a senior is at least about 60 years old. In certainembodiments, a senior is at about 70 years old. In certain embodiments,a senior is at least 80 years old.

In any of the embodiments provided in the present disclosure, thetopical, isotonic composition may be administered to an individualsubject as part of a treatment regimen, to members of a non-monadicsexual relationship (e.g., sexual dyad, sexual triad) as a part of atreatment regimen, or to both members of a non-sexual dyad. In certainembodiments, the sexual dyad is a homosexual dyad, a heterosexual dyad,or other sexual orientation dyad. In certain embodiments, a non-sexualdyad is parent-child dyad (e.g., mother-child or father-child),caregiver-child dyad, caregiver-adult patient dyad, or caregiver-seniorpatient dyad.

As discussed in HEIM Janneke et al., (2019) doi:10.1111/1471-0528.15870, hereby incorporated by reference in itsentirety, vaginal probiotics often involve the direct addition ofLactobaccilus strains to the urogenital and/or anogenital region, buthave significant drawbacks including regulatory barriers. The presentdisclosure is partially premised on compositions which are primarilyfocused on creating the optimal microbiotic environment for healthygrowth (e.g. Lactobaccilus growth) in the anogentical and/or urogenitalmicrobiomes. In some embodiments, the compositions may be used for thetreatment or prophylaxis of a disease, disorder, or condition associatedwith dysbiosis of the urogenital and/or anogenital region in a subjectin need thereof. The method for the treatment of the treatment orprophylaxis of a disease, disorder, or condition associated withdysbiosis of the urogenital and/or anogenital region in a subject inneed thereof may comprise administration of a composition of the presentdisclosure (e.g., a topical isotonic composition comprising bornylacetate, a prebiotic oligosaccharide, and a metal co-factor) to theurogenital and/or anogenital region of the subject. In some embodiments,the urogenital and/or anogenital region is the vagina. In someembodiments, the composition is administered topically to the vagina.The method may further comprise measuring the pH of the vagina (e.g.,with a pH nanosensor or other method) prior to application, andadministering a composition to effect the pH of the microbiomeenvironment in order to physiologically optimize Lactobacillusdominance. In various embodiments, the disease, disorder or conditionassociated with dysbiosis of the urogenital and/or anogenital region maybe selected from diabetes, lichen sclerosus, urinary incontinence,provoked vestibulodynia, vulvodynia, genital syndrome of menopause,interstitial cystitis, autoimmune genital disease, dyspareunia, orinfertility. In several embodiments, the compositions may decreasevaginal infection rates and/or urinary tract infections. In variousembodiments, the disease, disorder or condition associated withdysbiosis of the urogenital and/or anogenital region may be selectedfrom sexually transmitted diseases (e.g., HPV, HSV, HIV), cervicalcancer, pelvic floor disorder, genitourinary syndrome of menopause,provoked vestibulodynia, vulvodynia, interstitial cystitis, autoimmunegenital disease, dyspareunia, BV or infertility. In several embodiments,the compositions may decrease vaginal infection rates and/or urinarytract infections. In several embodiments, the compositions decreasepersistence of high risk HPV. Similarly, the anal microbiome has certaintolerances wherein the compositions can be used to promote healthygrowth therein. Measurements of the surface pH and human rectal mucosahas been measured in vitro to have a pH of from about 6.26 to about6.98, as shown in N McNeil, Gut 28 (1987): 707-713, hereby incorporatedby reference in its entirety. The rectum has been shown to have a morealkaline pH of 7.9 as shown in W Bitterman, et al., Gastroenterology 53(1967): 288-290, hereby incorporated by reference in its entirety. Insome embodiments, compositions for the treatment of the anal microbiome(e.g., lubricants) may have a pH of from about 5.5 to about 8 (e.g.,from about 5.5 to about 7, from about 5.8 and about 6.2).

In yet another aspect, the present disclosure provides a method forcollecting the genital microbiome from a donor dyad member comprisingtopically administering an effective amount of a topical, isotoniccomposition of the present disclosure to the urogenital and/oranogenital region of the donor dyad member and collecting the topical,isotonic composition from the urogenital and/or anogenital region of thesubject. In certain embodiments, the topical, isotonic composition iscollected in a receptacle. In certain embodiments, the topical, isotoniccomposition is administered via a wipe, adhesive roller, a blanket, anundergarment, diaper, film, or aerosol. In some embodiments, thecomposition is integrated into a tampon, vaginal ring, cervical cup,diaphragm, or condom, wherein the composition will be released uponinsertion into the urogenital and/or anogenital region.

In certain embodiments, the collected topical, isotonic composition isassayed for the presence of pathogenic microorganisms; cultured toidentify and propagate beneficial microorganisms (e.g., Lactobacilli);or both. In certain embodiments, the beneficial microorganisms areisolated and added as a probiotic or by vaginal flora transfer to aseparate, topical, isotonic composition of the present disclosure foradministration to a recipient dyad member or other unrelated individual.In certain embodiments, the donor dyad member and recipient dyad memberare members of a sexual dyad, e.g., a heterosexual dyad, homosexualdyad, or other sexual orientation dyad.

The individual components of the compositions described herein (e.g.,prebiotic oligosaccharides, metal co-factors, bornyl acetate, essentialoils comprising bornyl acetate) may be used for application of a subjectin need thereof. In some embodiments, these individual components (e.g.,prebiotic oligosaccharides, metal co-factors, bornyl acetate, essentialoils comprising bornyl acetate) may be used for the preparation of amedicament (e.g., topical compositions, isotonic compositions, topicalisotonic compositions) for the treatment of a subject in need thereof.For example, the individual components or the medicament may beadministered to the subject in order to hydrate the urogenital and/oranogenital region of the subject and/or lubricate the anogenital regionof the subject and/or clean the urogenital and/or anogenital region ofthe subject and/or decrease irritation or inflammation of the urogenitaland/or anogenital region of the subject and/or enhance the genitalmicrobiota of the subject. In certain embodiments, the individualcomponents and compositions described herein may be used for thetreatment or the prophylaxis of the dysbiosis of a subject in needthereof.

Applicators

The present disclosure also includes applicators which may be used foradministration to the urogenital and/or anogenital region of a subject(e.g., the vagina). As shown in FIG. 1, applicator 1 comprises storageportion 2 and delivery portions 14 and 15. Storage portion 2 comprisesan internal reservoir configured to house a composition (e.g., acomposition as disclosed herein for application to the urogenital and/oranogenital region). Storage portion 2 comprises bulb 4 with grippingelements 5 and connector 6 which connects storage portion 2 to deliveryportion 14. The connector portion may be configured to connect todelivery portion 14 through one or more removable connectors such asthreading on both elements, snap connections, and the like. Whenconnected, the internal reservoir is in fluid communication with aninternal flow element in delivery portions 14 and 15. Force applied tobulb 4 from a user causes composition within the internal reservoir toflow through delivery portions 14 and 15 and out exit orifice 16.Delivery portion 14 has a central axis 11 and delivery portion 15 has acentral axis 10. These two axes are angled with respect to one anotherat an angle 13 of, for example, less than about 90° or less than about60° or less than about 45° or less than 30° or less than about 20°. Insome embodiments, the angle between axis 10 and axis 11 is movable suchthat it may be set by a user. Delivery element 15 is dimensioned forinsertion into the urogenital and/or anogenital region of a user. Forexample, delivery element 15 may be substantially cylindrical about itsmajor axis 10 and comprises a rounded distal end at exit orifice 16. Insome embodiments, delivery element 15 comprises a stopper 17 whichindicates to a user the maximum depth of insertion for the device. Inthe embodiment depicted, stopper 17 comprises two annular ringssurrounding around delivery element 15 to prevent further insertion ofthe applicator. In some embodiments, the length of delivery element(i.e., the length along axis 10) between exit orifice 16 and stopper 17is less than about 10 cm or less than about 9 cm or less than about 8 cmor less than about 7 cm or less than about 6 cm or less than about 5 cm.In some embodiments, the maximum circumference of delivery element 15 isless than about 8 cm or about 7 cm or about 6 cm or less than about 5 cmor less than about 4 cm or less than about 3 cm. FIG. 2 illustrates theuse of applicator 1. A user 50 holds storage portion 2 in their hands51. Due to the multiple axes of in the delivery portions, deliveryportion 15 may be easily inserted for application of the composition tothe user, while delivery portion 14 is not. As can be seen, applicationof composition stored within storage portion 15 to the urogenital and/oranogenital region may occur with one handed insertion into theurogenital and/or anogenital region without having to remove clothes orspread patient legs. The internal reservoir may hold an amount ofcomposition for a single use or have multiple uses. In some embodiments,the internal reservoir may have a volume of from about 5 mL to about 60mL or about 10 mL to about 50 mL or about 15 mL to about 30 mL or about20 mL to about 25 mL.

Applicator 1 may further comprise a sensor capable of indicating the pHof the surrounding environment. For example, the applicator may comprisea litmus dye which, following insertion, is capable if visuallydisplaying pH information about the environment of the urogenital and/oranogental regions. In some embodiments, the applicator may comprisenanosensor 60 such as that disclosed in U.S. Pat. No. 10,436,745, herebyincorporated by reference in its entirety and particularly in relationto pH nanosensors. Typically, the nanosensor is capable of measuring thepH of the surrounding environment (e.g., the anogenital and/orurogenital region). Following measurement, the nanosensor may be capableof communicating 62 the pH measurement (e.g., with Bluetooth, radiofrequency identification) with an external device such as a laptop,tablet, computer, server, and/or smart phone. The nanosensor maytransmit the value of the measured pH or information relating to the pH.For example, the sensor may transmit a binary signal and/or a ternarysignal depending on user settings. The external device may be configuredto interpret, display, and track such measurements.

In some embodiments, applicator may have a display 61 operable connectedto the nanosensor. In the embodiment depicted, display 61 isillustrating a “+” symbol indicating that the pH is above a value (in abinary signal setup), or above a range (in a ternary signal setup). Insome embodiments, the display may show the pH measured from thenanosensor. In various embodiments, the sensor may show a binary displaysymbols (e.g., a “+” for a pH measurement below a certain level such asabout 4.8 and a “−” for a pH measurement above and/or equal to thatlevel such as about 4.8). In some embodiments, the display may beconfigured to display ternary display symbols (e.g., a “0” if themeasured pH is at a pH or within a certain pH range, a “−” if the pH isbelow that certain pH or pH range, and a “+” if the measured pH is abovethat certain pH or pH ranges). In some embodiments, the display isconfigured to display ternary symbols wherein the certain pH range isfrom about 4.5 to about 5.

Another applicator is illustrated in FIGS. 3A-C. Applicator 20 isillustrated in storage (FIG. 3A), nearly application (FIG. 3B), anddisposal (FIG. 3C) configurations. Applicator 20 comprises a container25 such as a bag or pouch comprising an internal reservoir 26 as definedby seams 29. The pouch is removably sealed with closure 28. Theapplicator 20 may comprise a composition 27 for application such as, forexample, a composition as disclosed herein. Applicator portion 22 (alsoreferred to herein as a delivery element) comprises a stopper 23, and aninternal flow chamber extending from a distal end of applicator portion22 to exit orifice 33. Applicator portion 22 may comprise a nanosensor24. In some embodiments, the applicator portion may comprise ameasurement device capable of transmitting pH information of anenvironment (e.g., litmus paper). In the storage configuration (FIG.3A), applicator portion 22 is contained within the internal reservoir. Atwistable cap is connected to a cut out 30 on container 25 allowing forsealing of the applicator during storage at interface 31. In theembodiment depicted, the cap 21 is disposed proximal to exit orifice 33of applicator portion 22. The applicator portion may be forcibly removedfrom the container 25 by, for example, supply a force to cap 21 in adirection away from internal reservoir 26. Such force may break theinterface 31 between cap 21 and cutout 30 allowing the applicator to beremoved from internal reservoir 26. In some embodiments, the lengthapplicator portion 22 (i.e., the length along the major axis) betweenexit orifice 33 and stopper 23 is less than about 10 cm or less thanabout 9 cm or less than about 8 cm or less than about 7 cm or less thanabout 6 cm or less than about 5 cm. In some embodiments, the maximumcircumference of applicator portion 22 is less than about 8 cm or about7 cm or about 6 cm or less than about 5 cm or less than about 4 cm orless than about 3 cm. The internal reservoir may hold an amount ofcomposition for a single use or have multiple uses. In some embodiments,the internal reservoir may have a volume of from about 5 mL to about 60mL or about 10 mL to about 50 mL or about 15 mL to about 30 mL or about20 mL to about 25 mL.

As can be seen in FIG. 3B, the applicator may be pulled out of thecontainer where the stopper 23 and cutout 30 are adapted to preventscomplete removal of applicator portion 22 and keeping exit orifice 32 influid communication with internal reservoir 26. FIG. 3B illustratesapplicator 20 with applicator portion 22 removed from internal reservoir26. Once in this configuration, applicator 20 may be placed in theapplication configuration by removal of cap 21, for example, by twistingthe cap to expose exit orifice 32. In the application configuration,composition 27 may be applied, for example, to the anogenital and/orurogenital region following appropriate insertion. In some embodiments,composition 27 may be expelled through exit orifice 32 by applying aforce to container 29 (e.g., by squeezing container 29). As can be seen,nanosensor 24 is capable of communicating 43 with an external device.Following application, applicator 20 may be converted into storage modeby placing applicator portion 22 back into internal reservoir 26 andfully sealing closure 28. In some embodiments, nanosensor 24 is capableof communicating with an external device in storage and/or disposalconfigurations. In some embodiments, nanosensor 24 may communicate witha display located on application portion 22.

Referring now to FIGS. 4A and B, applicator 30 comprises a container 31comprising an internal reservoir 32 capable of holding a composition. Insome embodiments, the composition may be a composition of the presentdisclosure. Container 31 comprises a support 33 with distal end 34 whichmay be configured to be placed within internal reservoir 32 at supportdistal end 35. Within support 33 is a flow chamber in fluidcommunication with internal reservoir 32 running from distal end 35 todistal end 37. The applicator portion 36 (also referred to herein as adelivery element) with exit orifice 37 which may comprise sensor 38(e.g., nanosensor 38) is dimensionally configured to be placed andremovably attached over support 33 on container 31 as shown in FIG. 4B.Applicator portion 36 may be placed and secured on support 30 rotation42 of applicator portion 36 with respect to container 31. Such securingof applicator portion 36 may occur via complimentary threading betweensupport 30 and applicator portion 36. The sensor 38 may be configured totransmit pH information 43 to an external device and/or send pHinformation to a display 44. In some embodiments, the length ofapplicator portion 36 (i.e., the length along the major axis) is lessthan about 10 cm or less than about 9 cm or less than about 8 cm or lessthan about 7 cm or less than about 6 cm or less than about 5 cm. In someembodiments, the maximum circumference of applicator portion 36 is lessthan about 8 cm or about 7 cm or about 6 cm or less than about 5 cm orless than about 4 cm or less than about 3 cm. The internal reservoir mayhold an amount of composition for a single use or have multiple uses. Insome embodiments, the internal reservoir may have a volume of from about5 mL to about 60 mL or about 10 mL to about 50 mL or about 15 mL toabout 30 mL or about 20 mL to about 25 mL.

In some embodiments, the compositions of the present disclosure may bedelivered using vaginal or topical films wherein the composition iscapable of diffusing from the film into its surrounding environment.Suitable film formers include chitosan, hydroxypropyl methylcelluloseand blends of these polymers (e.g., with 40% PEG 400 as plasticizer), apolymeric matrix/chitosan with carrageenan (κ-, λ-, and t-), pectin andgellan gum, hydroxyl propylcellulose and sodium alginate as polymers andpropylene glycol and polyethylene glycol-400 as plasticizers, polyvinylalcohol, poloxamer 407 and 188, hypromellose, sodiumcarboxymethylcellulose, hydroxylpropylmethylcellulose,hydroxyethylcellulose and polyvinyl pyrrolidone K-90, hydroxypropylmethylcellulose and Eudragit polymers (e.g., Eudragit RL100) andpropylene glycol as plasticizer, hydroxypropyl methylcellulose,polyvinyl alcohol, polyethylene oxide, glycerol,poly(2-oxazoline)/polyoxazoline polymers and combinations thereof.

The applicators (or any portion thereof such as the container and/or theapplicator element) may be formed from those materials known in the art.In some embodiments, portions of the applicator or the entire applicatormay be made low waste packaging materials such as biodegradableplastics. Suitable biodegradable plastics may be bio-based plastics suchas polyhydroxyalkanoates (PHAs), polylactic acid (PLA), starch blends,cellulose-based plastics, lignin-based polymer composites, andcombinations thereof. The biodegradable plastics may also be petroleumbased such as polyglycolic acid (PGA), polybutylene succinate (PBS),polycaprolactone (PCL), poly(vinyl alcohol) (PVA, PVOH), polybutyleneadipate terephthalate (PBAT), and combinations thereof. In someembodiments, portions of the applicator (e.g., the applicator element)are composed of paper and/or cardboard.

EXAMPLES Example 1: Foaming Wash & Genital Prebiotic Conditioner

Baseline pH levels in 6 circumcised white males ranging from 22-60 yrsof age were identified.

Left antecubital fossa samples were chosen as a control with no subjectshaving showered in the previous 10 hours or more prior to sampleacquisition. From these samples, a pH (average±SD) of 4.52±0.52 wasfound for these subjects.

Penile pH levels at several positions were measured. Average pH valueswere 4.96+/−0.52 (at the dorsal root); 5.30+/−0.52 (around thecircumference of the corona of the glans penis); and 5.24+/−0.52 (overthe external meatus). In 5 of the 6 men, penile pH was higher at themeatus than at the dorsal root. Penile pH levels, showed a gradient(P<0.05) towards a higher pH moving distally forward on the penis, withthe glans and coronal sulcus and external meatus being a higher pH thanthe distal root of the penis (and the forearm control). The average pHvalues and the standard deviation at each measured position are shown inFIG. 5.

The impact of prebiotic genital care systems on pH at these locationswas assessed using a penile foaming wash followed by a leave-onconditioning lubricant, applied to the penis (n=4). Leading commercialproducts, containing ingredients known to harm health microbiomespecies, were also measured. Specifically Nivea Men DEEP Active Clean(listed ingredients: Water, Sodium Laureth Sulfate, CocamidopropylBetaine, Acrylates Copolymer, PEG-7 Glyceryl Cocoate, Fragrance,Charcoal Powder, PEG-200 Hydrogenated Glyceryl Palmate, PEG-40Hydrogenated Castor Oil, PEG-3 Distearate, Trisodium EDTA, SodiumHydroxide, Phenoxyethanol, Methylparaben, Ethylparaben) followed byAstroglide lubricant (listed ingredients: Purified Water, Glycerin,Propylene Glycol, Polyquaternium 15, Methylparaben, Propylparaben) werecompared to formulations of the disclosure. In these experiments, thecomponents of the penile foaming wash and leave-on conditioninglubricant measured are shown in Table 8.

TABLE 8 Wash - Formula Conditioner - Formula Ingredient A (wt %) B (wt%) Purified Water, USP 94.847 98.972 Sodium chloride 0.100 0.300Lactulose (prebiotic 0.050 0.050 oligosaccharide) Rosmarinus officinalis0.050 essential oil (essential oil comprising bornyl acetate) Abiessibirica essential oil 0.020 (essential oil comprising bornyl acetate)Hypromellose 0.300 0.400 Sodium benzoate 0.250 Gluconolactone 0.750Arabinogalactan 0.100 Mentha spicata essential oil 0.020 0.015 (Biofilminhibiting agent) Citrus aurantium var. 0.010 0.010 amara essential oil(flavonoid) Manganese chloride (Metal 0.0025 0.0025 co-factor) SodiumCocyl Isethionate 1.000 Coco Betaine (36% active) 1.000 Citric Acid(buffering agent, QS to pH 5.0 pH adjusting agent) Bornyl acetate (notadded 0.010 from essential oils) Monosodium phosphate, 0.240 anhydrous(buffering agent) Sodium hydroxide (pH QS to pH 4.8 adjusting agent)

The impact on pH of of Formula A foaming wash and Formula B as a leaveon conditioner to the penis, were evaluated. Penile pH was measured atbaseline, and after three days of treatment with each product. Theresults are shown in FIG. 6.

Significant impacts of the treatment type on pH values were observed(p<0.05) for all measurements. The Nivea/Astroglide treatment over 3days significantly RAISED penile pH levels (Mean+/−SD) as compared tothe pH levels observed at baseline and after use of the prebioticformulations AB at the external meatus. The commercial products alsostrongly raised penile pH (approaching pH 6; p<0.01) over that found atthe unwashed antecubital fossa control.

Anerobic bacterial infections of the penis, especially from distalsites, have been associated with: vaginal infections in partners; andgreater risk of STDs and HIV in men. Elevated penile pH levels may beassociated with a reduction in healthy microbial species of the humangenital organs. Application of products that elevate skin pH tends todestabilize the normal protective glycocalyx of skin. Other studies haveshown an association between higher skin pH and disease. As can be seen,the compositions of the present disclosure have an effect of minimizingpenile pH change following their application.

Example 2: Formulations

Several wash and conditioning formulations have been prepared with thecomponents as shown in Tables 9 (wash) and 10 (conditioner).

TABLE 9 Form. Form. Form. a1 (% a2 (% a3 (% Ingredient Name by wt.) bywt.) by wt.) Purified Water, USP 95.645 91.227 94.108 Sodium chloride0.1000 0.200 0.100 Lactulose (prebiotic oligosaccharide) 0.0500 0.1000.750 Picea mariana essential oil 0.0200 Hydroxyethyl cellulose 0.4000Sodium dehydroacetate 0.1500 0.150 Arabinogalactan 0.1000 0.100 00Mentha spicata essential oil (biofilm 0.0200 0.020 0.020 inhibitingagent) Citrus paradisi essential oil 0.0100 (flavonoid) Manganesechloride (metal co-factor)* 0.005 0.0025 0.0015 Sodium Cocyl Isethionate1.500 1.250 1.500 Cocamidopropyl Betaine^(†) 2.000 6.300 1.000 CitricAcid (buffering agent, pH QS to QS to modifying agent) pH 5.0 pH 5.0Rosmarinus officinalis essential oil 0.050 0.050 (essential oilcomprising bornyl acetate) Cetyl hydroxyethylcellulose 0.600 Gycerolcaprylate 0.025 Sodium benzoate 0.100 Pseudotsuga menziesii essentialoil 0.015 Citrus aurantium var. amara essential 0.010 0.010 oil(flavonoid) Lactic Acid (buffering agent, pH QS to modifying agent) pH5.0 Abies sibirica essential oil (essential 0.01 oil comprising bornylacetate) Hydroxypropyl guar gum 0.300 Polysorbate 20 2.000 *Theindicated weight percentage of metallic co-factor (i.e., magnesiumchloride) was achieved by adding the appropriate amount of a hydrate ofthe metallic salt (e.g., magnesium chloride (II) tetrahydrate). ^(†)Theindicated weight percent of Cocamidopropyl betaine is the weight percentfrom a 36% solution of cocamidopropyl betaine added to each solution.

TABLE 10 Form. Form. Form. b1 (% b2 (% b3 (% Ingredient Name by wt.) bywt.) by wt.) Purified Water, USP 98.832 98.617 96.825 Monosodiumphosphate, anhydrous 0.240 Sodium chloride 0.300 0.500 0.500 Lactulose(prebiotic oligosaccharide) 0.050 0.10 0.050 Abies sibirica essentialoil (essential 0.020 0.020 oil comprising bornyl acetate) Hydroxyethylcellulose 0.400 Mentha spicata essential oil (biofilm 0.015 inhibitingagent) Sodium Benzoate 0.130 Citrus aurantium var. amara essential 0.0100.010 oil (flavonoid) Manganese chloride (metal co-factor)* 0.003 0.003Lactic Acid (buffering agent, pH QS to modifying agent) pH 4.8Rosmarinus officinalis essential oil 0.040 (essential oil comprisingbornyl acetate) Pseudotsuga menziesii essential oil 0.015 Hydroxypropylguar 0.550 Sodium dehydroacetate 0.150 Mentha aquatica essential oil(biofilm 0.015 inhibiting agent) Citric Acid (buffering agent, pH QS tomodifying agent) pH 4.8 Hypromellose 0.600 Lactobacillus Ferment 2.000Gluconolactone 0.005 Sodium hydroxide (pH adjusting agent) QS to pH 4.8*The indicated weight percentage of metallic co-factor (i.e., magnesiumchloride) was achieved by adding the appropriate amount of a hydrate ofthe metallic salt (e.g., magnesium chloride (II) tetrahydrate).

Example 3: Measurements on Post-Menopausal Women

Several vaginal wash and lubricant recipes of the present disclosure(Formula B2, Formula D 71519C) using either pure bornyl acetate or in anessential oil, when used in two post-menopausal women (ages 59 and 46),both undergoing hormone replacement therapy (HRT), have had thesurprising effect of substantially:

1) increasing arousal and decreased latency to orgasm as compared to notreatment or treatment with commercially available products such asAstroglide, Ky., and Replens; and

2) long term lubrication, hydration and comfort as compared to notreatment or treatment with commercially available products such asastroglide, KY, and replens.

Parameters were measured as disclosed in DL Rowland, et al., J Sex Med15 (2018): 1463-1471, A. Huang et al., Menopause 17 (2010): 121-126, E.Erekson et al., Menopause: The Journal of the North American MenopauseSociety 20 (2013): 973-979, B. Ettinger, Menopause 15 (2008): 889-889,E. Gerstenberger, et al., Jour of Sexual Med 7 (2010): 3096-3103, eachhereby incorporated by reference in their entirety.

Example 4: Effect of Compositions on Mouse In-Vitro Fertilization andEmbryo Development (IVF-MEA)

Two topical isotonic composition was prepared with the components shownin Table 11. Formula D6 had a total bornyl acetate content of 0.02% byweight of the composition.

TABLE 11 Formula D Formula D6 Ingredient Name (% by wt) (% by wt)Solvent 98.077 98.019 Buffering agent 0.240 0.483 Isotonicity agent0.300 0.100 Prebiotic oligosaccharide (combination 0.300 0.300 ofLactulose and gluconolactone) (−)-Bornyl acetate 0.005 Essential oilcomprising bornyl acetate 0.020 (Abies sibrica extract)Viscosity-increasing agent 1.000 1.000 Humectant 0.050 0.050 Biofilminhibiting agent 0.015 0.015 Flavanoid 0.010 0.010 Manganese chloride*0.003 0.003 pH adjusting agent QS to pH 4.5 QS to pH 4.5 *The indicatedweight percentage of metallic co-factor (i.e., magnesium chloride) wasachieved by adding the appropriate amount of a hydrate of the metallicsalt (e.g., magnesium chloride (II) tetrahydrate).

10% vol/vol concentrations of the compositions in Table 11 were preparedin in vitro fertilization (IVF) media. Mouse oocytes were placed withmouse sperm in formula D6 for 4 hours. After 4 hours of incubation at37° C. and 5.0% CO₂, 21 fertilized mouse oocytes were washed andtransferred to culture medium for 96 hours at 37° C. and 5.0% CO₂.Results following exposure to the 10% formulation solution are comparedto mouse ova placed with sperm and fertilized in a control culturemedium for 4 hours. After 4 hours of incubation at 37° C. and 5.0% CO₂,21 fertilized mouse oocytes were washed and transferred to culturemedium for 96 hours at 37° C. and 5.0% CO₂. Four-hour incubation withformula D6 resulted in 83% of oocytes fertilized, compared to 96% ofcontrol. 100% of oocytes exposed to D6 and to control developed toblastocysts.

To be considered non-toxic, the percent of oocytes fertilized within 4hours in the formulation group should be 80% or more of that found inthe control group, and the percent of oocytes developing to expandedblastocyst at 96 hours in the formulation group must not be less than80% of that found in the control group. The results for the 10%concentration for 4 hour exposure test are shown in Table 12. As can beseen, Formula D6 is non-toxic.

TABLE 12 Oocytes Oocytes to Oocytes to expanded fertilized 2-cell withinblastocyst within within 4 hours 24 hours 96 hours Control 96% 100% 100%Formula D6 83% (86% of 100% 100% control)

Example 5: Effect of Compositions on Mouse Embryo Development (MEA)

10% vol/vol concentrations of the compositions in Table 11 were preparedin M2 culture media. One-cell mouse embryos were placed in the formulaD6 for 30 minutes or in culture media alone (control) at 37° C. and 5.0%CO₂. The embryos were then then transferred to culture media andincubated for 96 hours at 37° C. and 5.0% CO₂. Results followingone-cell mouse embryo exposure to the 10% formulation solution arecompared one-cell mouse embryo in a control culture medium for 30minutes. 30 minute incubation resulted in 100% of the oocytes undergoingdivision within 24 hours and 100% of oocytes converting to expandedblastocyst within 96 hours for both control and test settings.

To be considered non-toxic, the number of oocytes developing to expandedblastocyst at 96 or 120 hours in the formulation group must not be lessthan 80% of that found in the control group. The results for the 10%concentration for 4 hour exposure test are shown in Table 13. As can beseen, Formula D6 is non-toxic.

TABLE 13 1-cell embryos to 1-cell embryos to 2-cell embryos expandedblastocyst within 24 hours within 96 hours Control 100% 100% Formula D6100% 100%

Example 6: Human Sperm Motility Measurements

A semen sample was obtained from a healthy normospermic donor. Thespecimen was produced by masturbation without lubricant into a sterileplastic container after a recommended abstinence period of 48-96 hours.The specimen was allowed to liquefy and then processed within 30minutes. Spermatozoa were separated from liquefied semen by densitygradient separation. The washed sperm were then resuspended in spermwash medium. Formula D was added to an aliquot of the sperm sample toachieve a 10% V/V concentration. An aliquot from the same sperm samplebut without Formula D6, serves as the medium-only control. The sampleswere incubated in the same incubator at 32° C. and 5% CO₂ for 24 hours.Sperm motility following exposure to the test-item for 30 minutes mustbe ≥80% of control to be considered non-toxic. The sperm motilitymeasurements initially and after 24-hours are shown in Table 14.

TABLE 14 Initial Motility 24-hour motility Control 97% 97% Formula D697% (100% of 97% (100% of control) control)

Example 7: Bovine Sperm Motility Studies

Commercial cryopreserved sperm from 2 bulls was pooled and used in allreplicates. Semen straws (0.5 mL) were thawed and sperm concentrationwas adjusted to 10 million spermatozoa per mL using a commercial bovinesemen-freezing medium.

Straws containing sperm were thawed for use in studies, where they weremixed with 10% gel treatment with one of the gel compositions in Table15 and incubated for 30 min at 39° C. in a CO₂ incubator. The bornylacetate content (coming from essential oils comprising bornyl acetate)for each formulation is also provided in Table 15.

TABLE 15 71519A 71519B 71519C 0618CA45 0618CA68 (% by wt) (% by wt) (%by wt) (% by wt) (% by wt) Total bornyl acetate 0.045 0.025 0.055 0.020.02 content Ingredient Name Purified water, USP 97.344 97.264 97.23497.739 97.739 Disodium phosphate 0.531 0.531 0.531 0.531 0.531 Lacticacid 0.500 0.500 0.500 Sodium chloride 0.100 0.100 0.100 0.100 0.100Lactulose 0.0500 0.050 0.050 0.050 0.050 Abies sibirica 0.0250 0.0250.025 0.020 0.020 Hypromellose 0.500 0.500 0.500 0.400 0.400Gluconolactone 0.500 0.500 0.500 0.100 0.100 Hydroxypropyl guar 0.4000.400 0.400 0.400 0.400 gum Rosmarinus officinalis 0.020 0.020 Menthaspicata 0.010 0.010 0.010 0.008 0.008 Citrus aurantium var. 0.015 0.0150.015 Amara Manganese chloride* 0.005 0.005 0.005 0.00455 0.0054Arabinogalactan 0.100 0.100 0.200 0.200 Picea mariana 0.010 Sodiumhydroxide QS to QS to QS to QS to QS to pH 4.8 pH 4.8 pH 4.8 pH 4.8 pH6.8 Citric acid 0.409 0.409 Oleuropein 0.0200 0.0200 Citrus reticulata0.0100 0.0100 Juniperus communis 0.0080 0.0080 *The indicated weightpercentage of metallic co-factor (i.e., magnesium chloride) was achievedby adding the appropriate amount of a hydrate of the metallic salt(e.g., magnesium chloride (II) tetrahydrate).

After incubation and thorough mixing of samples, replicate aliquots wereremoved from treatments for computer assisted sperm analysis (CASA)using a Hamilton Thorne IVOS analyzer. Briefly, for each replicate, a 5μL sample is loaded on a MicroCell sperm counting chamber. The CASAstage is set at 39° C. for analysis. A minimum of 5 fields per sampleare analyzed for measurement of sperm motility and concentration.Analysis includes % motile sperm, % progressively motile sperm, andtotal motile sperm concentration, as well as numerous measurements ofsperm motion including velocity and lateral head displacement. Spermmotility parameters following exposure to the test-item for 30 minutesmust be ≥80% of control.

Generally, motility at 80% or greater is viewed as acceptable by theFDA. Total percent of motile sperm at 10 and 30 min and total percentprogressively motile sperm at 10 and 30% were determined. Results fortotal percent motile sperm are shown in FIG. 7A and total percent forprogressively motile sperm are shown in FIG. 7B. As can be seen, theaddition of citric acid at pH 4.5 or 4.8 had a significantly negativedeleterious impact on sperm function. With this exception, all formulassupported high levels of sperm total motility and progressive motility,with some formulas exhibiting sperm function equal to or greater thanthat of controls.

Formulations with a pH of 6.8 were also prepared with the components asshown in Table 16 and motility measurements were performed. The motilitymeasurement results for these compositions are shown in FIGS. 8A and 8B.

TABLE 16 060919C 0617HF 0617HL 0617HM Ingredient Name (wt %) (wt %) (wt%) (wt %) Purified Water, USP 96.9744 97.109 97.0285 97.1235hypromellose 0.6 0.6 0.6 0.6 Disodium phosphate, 0.531 0.531 0.531 0.531anhydrous gluconolactone 0.5 0.56 0.56 0.56 lactic acid 0.5 0.5 0.5 0.5Hydroxypropyl guar gum 0.4 0.4 0.4 0.4 Arabinogalactan 0.2 0.2 0.2 0.2Sodium hydroxide 0.19 0.206 0.16 0.208 Sodium chloride 0.1 0.05 0.050.05 Lactulose 0.05 0.0025 0.1 0.0025 Abies sibirica 0.02 0.025 0.0080.008 Citrus paradisi 0.01 0.01 0.01 0.01 Mentha spicata 0.01 0.01 0.010.01 Manganese chloride* 0.0046 0.0025 0.0025 0.005 pH 6.8 6.8 6.8 6.8*The indicated weight percentage of metallic co-factor (i.e., magnesiumchloride) was achieved by adding the appropriate amount of a hydrate ofthe metallic salt (e.g., magnesium chloride (II) tetrahydrate).

Motility measurements were also performed on the compositions shown inTable 17.

TABLE 17 1004BA5 1004BA10 1004BA15 Ingredient Name % by wt. % by wt. %by wt. Purified Water, USP 98.077 98.072 98.067 Monosodium phosphate,0.240 0.240 0.240 anhydrous Sodium chloride 0.300 0.300 0.300 Lactulose0.050 0.050 0.050 Bornyl acetate 0.005 0.010 0.015 Hypromellose 0.6000.600 0.600 Gluconolactone 0.250 0.250 0.250 Hydroxypropyl guar gum0.400 0.400 0.400 Arabinogalactan 0.050 0.050 0.050 Mentha spicata 0.0150.015 0.015 Neroli oil 0.010 0.010 0.010 Manganese chloride* 0.003 0.0030.003 Sodium hydroxide QS to pH 4.5 QS to pH 4.5 QS to pH 4.5 *Theindicated weight percentage of metallic co-factor (i.e., magnesiumchloride) was achieved by adding the appropriate amount of a hydrate ofthe metallic salt (e.g., magnesium chloride (II) tetrahydrate).

The compositions disclosed in Table 17 were measured to have effect onsperm motility as shown in Table 18.

TABLE 18 1004BA5 1004BA1 1004BA15 % of control % of control % of controlTotal Motile (10 min) 109 89 98 Total Motile (30 min.) 90 87 97Progressively Motile 109 82 93 (10 min.) Progressively Motile 97 85 100(30 min.)

Example 8: Formulations

Several compositions were formulated with the components as shown inTable 19.

TABLE 19 Formula Da Formula Db Formula Dd Ingredient Name (% by wt) (%by wt) (% by wt) Purified water, USP 97.576 97.274 98.067 Monosodiumphosphate, 0.523 0.240 anhydrous Disodium phosphate 0.531 0.325 Lacticacid 0.500 Sodium chloride 0.200 0.100 0.300 Lactulose 0.050 0.050 0.050Abies sibirica 0.015 Xanthan gum 0.900 Mentha spicata 0.015 0.050 0.015Rosmarinus officinalis 0.010 Sodium dehydroacetate 0.200 Manganesechloride* 0.003 0.003 0.003 Sodium hydroxide QS to QS to QS to pH 4.5 pH4.5 pH 4.5 Pseudotsuga menziesii 0.01 Kappa-Carrageenan 1.000Gluconolactone 0.250 Cetyl hydroxyethylcellulose 0.40 Citrus aurantiumvar. 0.015 0.010 Amara Picea mariana 0.015 Hydroxyethyl cellulose 1.100Arabinogalactan 0.050 Sodium benzoate 0.15 *The indicated weightpercentage of metallic co-factor (i.e., magnesium chloride) was achievedby adding the appropriate amount of a hydrate of the metallic salt(e.g., magnesium chloride (II) tetrahydrate).

Example 9: Safety Testing of Lubricants

Lubricant formulations were prepared with varying concentrations of(−)-bornyl acetate. The formulations are shown in Table 20.

TABLE 20 Ingredient Name % by wt. % by wt. % by wt. Purified Water, USP98.077 98.072 98.067 Monosodium phosphate, 0.240 0.240 0.240 anhydrousSodium chloride 0.300 0.300 0.300 Lactulose 0.050 0.050 0.050 Bornylacetate 0.005 0.010 0.015 Hypromellose 0.600 0.600 0.600 Gluconolactone0.250 0.250 0.250 Hydroxypropyl guar gum 0.400 0.400 0.400Arabinogalactan 0.050 0.050 0.050 Mentha spicata 0.015 0.015 0.015Neroli oil 0.010 0.010 0.010 Manganese chloride* 0.003 0.003 0.003Sodium hydroxide QS to QS to QS to pH 4.5 pH 4.5 pH 4.5 *The indicatedweight percentage of metallic co-factor (i.e., magnesium chloride) wasachieved by adding the appropriate amount of a hydrate of the metallicsalt (e.g., magnesium chloride (II) tetrahydrate).

Each lubricant was used during the intercourse of a sexual dyad. Noadverse effects were observed aside from the composition comprisingpurified 0.015% (−)-bornyl acetate by weight which resulted inshort-term mild mucosal irritation in a member of the sexual dyad onoccasion.

Example 10: Muco-Adhesion Potential of the Compositions

A formulation was prepared with the components as shown in Table 21.

TABLE 21 Formula PL1116SSB Ingredient Name % by wt. Purified Water, USP97.872 monosodium phosphate, anhydrous 0.240 sodium chloride 0.300Lactulose 0.050 (−)-Bornyl Acetate 0.010 hypromellose 0.600gluconolactone 0.250 hydroxypropyl guar gum 0.400 Arabinogalactan 0.050Mentha spicata (spearmint) 0.015 Neroli oil 0.010 Manganese chloride*0.003 Sodium Benzoate 0.125 Sodium Salicylate 0.075 Sodium Hydroxide QSto pH 4.5 *The indicated weight percentage of metallic co-factor (i.e.,magnesium chloride) was achieved by adding the appropriate amount of ahydrate of the metallic salt (e.g., magnesium chloride (II)tetrahydrate).

Mucoadhesion potential of the formulation and two competitive productswas determined in vitro using solutions of natural porcine gastricmucin. Properties and behavior of gastric mucin are similar (if notidentical) to mucosa from other regions such as the mucosa of theurogenital and/or anogential regions. Muco-adhesion occurs when twocomponents, (one being biological in origin), are held together byinterfacial forces, including electrical charge and polymer chainentanglement. Muco-adhesive properties may allow better contact of eachformulation with the vaginal surface. Superior muco-adhesion potentialsuggests the composition is better able to be retained in the vaginathus aiding in supplementation of vaginal secretions and moisture. Also,if used for drug delivery compositions with increased muco-adhesivepotential would have superiority with regard to residence time anddelivery of active agent.

The level of muco-adhesion for several compositions was quantified bymeasuring the change in viscosity due to synergism in a mucin-vaginalmoisturizer gel solution as described in D Ivarsson, et al., Colloidsand Surfaces B 92 (2012): 353-359, hereby incorporated by reference inits entirety and particularly in relation to the mucin-vaginalmoisturizer gel solution. Mucoadhesion strength of compositions wasassessed by co-mixing each test composition with 8% mucin from porcinestomach Type II (Sigma-Aldrich). The mucin solution was prepared with0.9% NaCl solution (saline). The 8% mucin solution was mixed at roomtemperature with one of the test lubricants (Replens, RepHresh,PL1116SSB Formulation of Table 21) to provide final concentrations of25% (wt/wt) gel/mucin solutions. The Replens lubricant comprisedpolycarbophil, mineral oil, hydrogenated palm oil glyceride, glycerin,carbomer homopolymer type B, sodium hydroxide and sorbic acid and had apH of 2.95. The RepHresh lubricant comprised Purified Water USP,Glycerin, Polycarbophil, Carbopol 974P, Ethylparaben Sodium,Methylparaben Sodium, Propylparaben Sodium, and Sodium Hydroxide and hada pH of 3.46. The relative viscosity of the 8% mucin-only solution wascompared to the relative viscosity of the three-resulting mucin-gelsolutions by measuring the time required for a 1 mL drop oflubricant/mucin solution to travel 90 mm on an uncoated cardboardsurface oriented at a 45° angle. The experiment was performed induplicate.

Mucoadhesion differed by gel type. RepHresh showed no muco-adhesion, asdemonstrated by no change in solution viscosity compared to control(p=0.92). Replens showed a small, but significant (p=0.04) increase ofapproximately 2.75-fold time required to travel the 90 mm, compared tocontrol (5.5 seconds vs 2 seconds). In contrast, the PL1116SSBformulation of the present disclosure showed an 8.5 fold increase(p<0.001) in relative viscosity versus control (17 seconds versus 2seconds). suggesting a high mucoadhesion potential for the compositionsof the present disclosure (see FIG. 9. The viscosity differences cannotbe explained based on differences in the viscosities of the neat gels.Replens is approximately 5 times, and RepHresh 20 times more viscousthan the PL1116SSB Formulation.

Specific Embodiments

Non-limiting specific embodiments are described below each of which isconsidered to be within the present disclosure.

Specific Embodiment 1. A topical, isotonic composition comprising:

(a) a prebiotic oligosaccharide in an amount ranging from about 0.001%to about 5% by weight of the composition;

(b) a metal co-factor in an amount ranging from about 0.0001% to about0.1% by weight of the composition; and

(c) borneol, pharmaceutically acceptable salts thereof, or prodrugs(e.g., ester prodrug such as bornyl acetate) of any of the foregoing inan amount ranging from about 0.00025% to about 0.15% by weight of thecomposition

wherein the pH of the composition ranges from about 3.5 to about 8.

Specific Embodiment 2. The topical, isotonic composition of SpecificEmbodiment 1, wherein the prebiotic oligosaccharide comprises raffinose,lactulose, trehalose, galactooligosaccharide, alpha-glucanoligosaccharide, beta-glucan oligosaccharide, fructooligosaccharide,isomaltooligosaccharide, maltose, inulin, pectin, or any combinationthereof.

Specific Embodiment 3. The topical, isotonic composition of SpecificEmbodiment 1 or 2, wherein the metal co-factor comprises zinc, selenium,manganese, cobalt, iron, copper, or any combination thereof.

Specific Embodiment 4. The topical, isotonic composition of any one ofSpecific Embodiments 1-3, wherein the composition comprises bornylacetate in an amount ranging from about 0.00025% to about 0.15% byweight of the composition.

Specific Embodiment 5. The topical, isotonic composition of claim 4,wherein the composition comprises from about 0.005% to about 0.5% of anessential oil comprising said bornyl acetate by weight of the essentialoil.

Specific Embodiment 6. The topical composition according to any one ofSpecific Embodiments 1-5 wherein the essential oil comprising bornylacetate comprises essential oil from Juniperus communis, Juniperusoccidentalis, Juniperus scopulorum, Abies sibirica, Abies alba, Abiesbalsamea, Abies fraseri, Abies grandis, Abies spectabilis, Abieskoreana, Abies procera, Abies nordmanniana, Abies magnifica, Abiespinsapo, Abies lasiocarpa, Abies concolor, Pseudotsuga menziesii,Rosmarinus officinalis, Ambrosia trifida, Pinus Mugo, Romanian solidago,Ribes nigrum, Laurus nobilis, or any combination thereof.

Specific Embodiment 7. The topical isotonic composition of any one ofSpecific Embodiments 1-6, further comprises a sesquiterpene alcoholand/or a monocyclic sesquiterpene.

Specific Embodiment 8. The topical isotonic composition according toSpecific Embodiment 7, wherein the sesquiterpene alcohol or monocycliccomprises nerolidol, an essential oil from Citrus aurantum (e.g., Citrusaurantium var. amata) and/or Citrus bigaradia, bisabolol, patchoulol,alpha-santalol, zingiberene, or combinations thereof.

Specific Embodiment 9. The topical, isotonic composition of SpecificEmbodiment 8, wherein composition comprises bisabolene in an amountranging from about 0.0001% to about 0.1% by weight of the composition.

Specific Embodiment 10. The topical, isotonic composition of any one ofSpecific Embodiments 1-9, further comprising a flavonoid in an amountranging from about 0.001% to about 0.1% by weight of the composition.

Specific Embodiment 11. The topical, isotonic composition of SpecificEmbodiment 10, wherein the flavonoid comprises catechin, epicatechin,rutin, luteolin, apigenin, kaempherol, myricetin, quercetin, quercitrin,naringin, naringenin, hesperetin, hesperidin, taxifolin, genistin,genistein, daidzein, cyanidin, apigenidin, tangeritin, fisetin,galangin, isorhamnetin, pachypodol, rhamnazin, pyranoflavonol,fluranoflavonol, eriodictyol, homoeriodictyol, taxifolin, gallocatechin,catechin 3-gallate, gallocatechin 3-gallate, epigallocatechin,epicatechin 3-gallate, epigallocatechin 3-gallate, theaflavin,thearubigin, proanthocyanidin, dephinidin, malvidin, pelargonidin,peonidin, petunidin, isoflavone, glycitein, isoflavane, isoflavandiol,isoflavene, coumestan, pterocarpan, myricitrin, phloridzin, or anycombination thereof.

Specific Embodiment 12. The topical, isotonic composition of any one ofSpecific Embodiments 1-11, further comprising a biofilm inhibiting agentin an amount ranging from about 0.001% to about 0.16% by weight of thecomposition.

Specific Embodiment 13. The topical, isotonic composition of SpecificEmbodiment 12, wherein the biofilm inhibiting agent comprises Lamiaceaeessential oil, Garcinia extract, Eurycoma longifolia extract, or anycombination thereof.

Specific Embodiment 14. The topical, isotonic composition of SpecificEmbodiment 13, wherein the Lamiaceae essential oil comprises essentialoil from Mentha spicata, Mentha pulegium, Mentha piperita, Menthaaquatica, Mentha arvensis, Mentha asiatica, Mentha australis, Menthacanadensis, Mentha cervina, Mentha citrata, Mentha crispata, Menthadahurica, Mentha diemenica, Mentha laxiflora, Mentha, longifolia, Mentharequienii, Mentha sachalinensis, Mentha satureioides, Menthasuavenolens, Mentha vagans, Melissa officinalis, Monarda Fistulosa, orany combination thereof.

Specific Embodiment 15. The topical, isotonic composition of SpecificEmbodiment 13 or 14, wherein the Garcinia extract comprises extract fromGarcinia mangostana, Garcinia travancorica, Garcinia cambogia, Garciniakola, Garcinia zeylanica, Garcinia xanthochymus, or any combinationthereof.

Specific Embodiment 16. The topical, isotonic composition of SpecificEmbodiment 13, wherein the Garcinia extract comprises a xanthoneselected from y-mangostin, garcinone-D, gartanin, and smeathxanthone.

Specific Embodiment 17. The topical isotonic composition of any one ofSpecific Embodiments 1-16, wherein the composition further comprises aprebiotic spice extract in an amount ranging from about 0.001% to about0.02% by weight of the composition.

Specific Embodiment 18. The topical, isotonic composition of SpecificEmbodiment 17, wherein the prebiotic spice extract comprises extractfrom ginger, black pepper, cayenne pepper, cinnamon, oregano, rosemary,turmeric, or any combination thereof.

Specific Embodiment 19. The topical, isotonic composition of any one ofSpecific Embodiments 1-18, further comprising a phytoesterol in anamount ranging from about 0.001% to about 0.1% by weight of thecomposition.

Specific Embodiment 20. The topical, isotonic composition of SpecificEmbodiment 19, wherein the phytoesterol comprises apigenin,β-sitosterol, campesterol, brassicasterol, stigmasterol, sitosterol, orany combination thereof.

Specific Embodiment 21. The topical, isotonic composition of any one ofSpecific Embodiments 1-20, further comprising a viscosity increasingagent in an amount ranging from about 0.1% to about 10% by weight of thecomposition.

Specific Embodiment 22. The topical, isotonic composition of SpecificEmbodiment 21, wherein the viscosity enhancing agent comprises guar gum,methylcellulose, ethylcellulose, ethyl methyl cellulose, ethylhydroxyethyl cellulose, carboxymethylcellulose, hydroxypropylcellulose,hydroxyethyl methyl cellulose, hydroxypropylmethylcellulose(hypromellose), hydroxyethylcellulose, cetyl hydroxyethycellulose,hydroxypropyl guar gum, glycosaminoglycans (e.g., hyaluronic acid),nonionic triblock copolymers such as poloxamers, gelatins, alginates,carrageenan, and agar, or any combination thereof.

Specific Embodiment 23. The topical, isotonic composition of any one ofSpecific Embodiments 1-22, further comprising a pH modifying agent in anamount ranging from about 0.01% to about 1% by weight of thecomposition.

Specific Embodiment 24. The topical, isotonic composition of SpecificEmbodiment 23, wherein the pH modifying agent comprises citric acid,lactic acid, formic acid, acetic acid, propionic acid, butyric acid,caproic acid, oxalic acid, maleic acid, benzoic acid, carbonic acid,ammonia, ammonium carbonate, diethanolamine, monoethanolamine, potassiumhydroxide, potassium phosphate dibasic, sodium bicarbonate, sodiumborate, sodium carbonate, sodium hydroxide, sodium lactate, sodiumphosphate dibasic, trolamine, or any combination thereof.

Specific Embodiment 25. The topical isotonic composition of any one ofSpecific Embodiments 1-24, further comprising a buffering agent in anamount ranging from about 0.01% to about 0.9% by weight of thecomposition.

Specific Embodiment 26. The topical, isotonic composition of SpecificEmbodiment 25, wherein the buffering agent comprises lactic acid andsodium lactate, citric acid and disodium phosphate, gluconolactone,citric acid and monosodium phosphate, or citric acid, monosodiumphosphate, and disodium phosphate.

Specific Embodiment 27. The topical, isotonic composition of any one ofSpecific Embodiments 1-26, wherein said composition is isotonic withphysiological fluid of the genital skin and/or mucosa.

Specific Embodiment 28. The topical, isotonic composition of any one ofSpecific Embodiments 1-27, wherein said composition is isotonic withphysiological genital skin barrier function.

Specific Embodiment 29. The topical, isotonic composition of SpecificEmbodiments 28, wherein said physiological genital fluid is selectedfrom cervico-vaginal fluids, urethral secretion, semen, or smega.

Specific Embodiment 30. The topical, isotonic composition of any one ofSpecific Embodiments 1-29 further comprising an osmolality adjustingagent, wherein the topical isotonic composition has an osmolality ofabout 120 mOsm/kg to about 450 mOsm/kg.

Specific Embodiment 31. The topical, isotonic composition of any one ofSpecific Embodiments 1-30, wherein the topical isotonic composition hasan osmolality of about 150 mOsm/kg or about 240 mOsm/kg.

Specific Embodiment 32. The topical, isotonic composition of SpecificEmbodiment 30 or 31, wherein the osmolality adjusting agent comprisesglucose, sucrose, sodium chloride, potassium chloride, calcium chloride,sodium sulfate, magnesium chloride, dextrose, mannitol, or anycombination thereof.

Specific Embodiment 33. The topical, isotonic composition of any one ofSpecific Embodiments 1-32, further comprising a preservative in anamount of about 0.001% to about 4% by weight of the composition.

Specific Embodiment 34. The topical, isotonic composition of SpecificEmbodiment 33, wherein the preservative comprises caprylyl glycol,cranberry extract, dichlorobenzyl alcohol, gluconolactone, green teaextract, oleuropein, pentylene glycol, phenethyl alcohol, pomegranateextract, benzoic acid, benzyl alcohol, dihydroacetic acid, ethylhexylglycerin, Lactobacillus ferment, pentylene glycol, potassium sorbate,sodium benzoate, sodium dehydroacetate, sodium salicylate, potassiumbenzoate, propanediol, resveratrol, hydantoin, or any combinationthereof.

Specific Embodiment 35. The topical, isotonic composition of any one ofSpecific Embodiments 1-34, wherein the topical, isotonic compositioncomprises from about 88% to about 98% solvent by weight of thecomposition, and the solvent comprises water.

Specific Embodiment 36. The topical, isotonic composition of any one ofSpecific Embodiments 1-35, further comprising a drug, an anogenitaland/or urogenital probiotic bacterial species, or both.

Specific Embodiment 37. The topical, isotonic composition of SpecificEmbodiment 36, wherein the drug is a hormone, an antiviral agent, anantifungal agent, an anti-inflammatory, an antibiotic, an erectiledysfunction treatment drug, a vaccine, a therapy for genitourinarysyndrome of menopause, an active ingredient for the treatment of lichensclerosus, an active ingredient for the treatment of interstitialcystitis, an active ingredient for the treatment of urinaryincontinence, or any combination thereof.

Specific Embodiment 38. The topical, isotonic composition of any one ofSpecific Embodiments 1-37, wherein the pH of the composition is about3.5 to about 6.8.

Specific Embodiment 39. The topical, isotonic composition of SpecificEmbodiment 38, wherein the pH of the composition is about 4.5.

Specific Embodiment 40. The topical, isotonic composition of any one ofSpecific Embodiments 1-37, wherein the pH of the composition is about 6to about 8.

Specific Embodiment 41. The topical, isotonic composition of SpecificEmbodiment 40, wherein the pH of the composition is about 6.8.

Specific Embodiment 42. The topical, isotonic composition of any one ofSpecific Embodiments 1-41, wherein the composition:

(1) hydrates the urogenital and/or anogenital region;

(2) lubricates the urogenital and/or anogenital region;

(3) cleans the urogenital and/or anogenital region;

(4) decreases irritation and/or inflammation of the urogenital and/oranogenital region;

(5) maintains and/or supports and/or enhances healthy genital andreproductive tract microbiota;

(6) supports and/or enhances genital tissue function;

(7) supports and/or enhancing genital fluid function;

(8) supports and/or enhances sperm viability and/or function;

(9) corrects and/or restores optimal genital pH;

(10) decreases bothersome genital symptoms;

or any combination thereof.

Specific Embodiment 43. The topical, isotonic composition of any one ofSpecific Embodiments 1-42, wherein the composition is formulated as aliquid, semi-solid, gel, jelly, film, foam, cream, douche, ointment,lotion, spray, suspension, emulsion, aerosol, suppository, particlesuspension, capsule, soap, suspension, emulsion, suppository, or paste.

Specific Embodiment 44. A method of hydrating the anogenital and/orurogenital region of a subject comprising topically administering aneffective amount of the topical, isotonic composition of any one ofSpecific Embodiments 1-43 to the anogenital and/or urogenital region ofthe subject.

Specific Embodiment 45. A method of lubricating the anogenital and/orurogenital region of a subject comprising topically administering aneffective amount of the topical, isotonic composition of any one ofSpecific Embodiments 1-43 to the anogenital and/or urogenital region ofthe subject.

Specific Embodiment 46. A method of cleaning the anogenital and/orurogenital region of a subject comprising topically administering aneffective amount of the topical, isotonic composition of any one ofSpecific Embodiments 1-43 to the anogenital and/or urogenital region ofthe subject.

Specific Embodiment 47. A method of decreasing irritation orinflammation of the anogenital and/or urogenital region of a subjectcomprising topically administering an effective amount of the topical,isotonic composition of any one of Specific Embodiments 1-43 to theanogenital and/or urogenital region of the subject.

Specific Embodiment 48. A method of enhancing the genital microbiota(e.g., beneficial genital microbiota such as Lactobacillus species) of asubject (e.g. prebiotic) comprising topically administering an effectiveamount of the topical, isotonic composition of any one of SpecificEmbodiments 1-43 to the urogenital and/or anogenital region of thesubject.

Specific Embodiment 49. The method of any one of Specific Embodiments44-48, wherein the subject is a human male.

Specific Embodiment 50. The method of any one of Specific Embodiments44-48, wherein the subject is a human female.

Specific Embodiment 51. The method of any one of Specific Embodiments44-48, wherein the subject is an intersex individual or a non-binarygendered individual.

Specific Embodiment 52. The method of any one of Specific Embodiments44-48, wherein the subject is undergoing gender reassignment and/or hasundergone gender reassignment.

Specific Embodiment 53. The method of any one of Specific Embodiments44-48, wherein said application occurs during masturbation of thesubject.

Specific Embodiment 54. The method of any one of Specific Embodiments44-53, wherein the topical, isotonic composition has a pH of about 3.5to about 6.8, and the subject is a human adult of about 12 years of ageor above.

Specific Embodiment 55. The method of Specific Embodiment 54, whereinthe human adult is a member of a non-monadic sexual relationship.

Specific Embodiment 56. The method of Specific Embodiment 55, whereinthe non-monadic sexual relationship comprises a sexual dyad or a sexualtriad.

Specific Embodiment 57. The method of Specific Embodiment 56, whereinthe topical, isotonic composition is administered to both members of thesexual dyad, and wherein both members of the sexual dyad are pubescentor older.

Specific Embodiment 58. The method of any one of Specific Embodiments48-57, wherein the topical, isotonic composition is administered priorto, during, or after sexual activity.

Specific Embodiment 59. The method of any one of Specific Embodiments48-58, wherein the topical, isotonic composition has a pH of about 6 toabout 8, and the subject is an infant of 0 to about 12 months of age, anadult of about 18 years to about 50 years of age who is actively tryingto conceive, or a senior of at least about 60 years of age.

Specific Embodiment 60. The method of Specific Embodiment 59, whereinthe subject is an adult of about 18 years to about 50 years of age whois actively trying to conceive is above the age of 50(e.g., is above theage of 60, is above the age of 70, is above the age of 80, is above theage of 90, is above the age of 100), and is a member of a non-monadicsexual relationship.

Specific Embodiment 61. The method of Specific Embodiment 60 wherein thenon-monadic sexual relationship comprises a sexual dyad or a sexualtriad.

Specific Embodiment 62. The method of Specific Embodiment 61, whereinthe topical, isotonic composition is administered to both members of thesexual dyad.

Specific Embodiment 63. The method of any one of Specific Embodiments59-62, wherein the topical, isotonic composition is administered priorto, during, or after sexual activity.

Specific Embodiment 64. The topical, isotonic composition of any one ofSpecific Embodiments 1-40, wherein the composition is administered via aroller ball, wipe, a blanket, an undergarment, an aerosol, a film, or adiaper.

Specific Embodiment 65. A device for implantation and/or insertion intothe anogenital and/or urogenital region of a subject; wherein the deviceis configured to release the composition according to any one of claims1-40; and said device is a tampon, vaginal ring, cervical cup,diaphragm, or condom.

Specific Embodiment 66. A method of hydrating, conditioning, cleansingor lubricating the urogenital and/or anogenital region of both membersof a sexual dyad, wherein the sexual dyad comprises at least one femalesubject, comprising:

(a) administering an effective amount of the topical, isotoniccomposition of any one of Specific Embodiments 1-43 to the urogenitaland/or anogenital region of both members of the sexual dyad during thenon-fertile phase of the female subject, wherein the topical, isotoniccomposition administered during the non-fertile phase has a pH of about4.5 and a tonicity of about 150 mOsm/kg.

(b) administering an effective amount of the topical, isotoniccomposition of any one of Specific Embodiments 1-43 to the urogenitaland/or anogenital region of both members of the sexual dyad during thefertile phase of the female subject, wherein the topical, isotoniccomposition administered during the fertile phase has a pH of about 6.8and a tonicity of about 240 mOsm/kg.

Specific Embodiment 67. The method of Specific Embodiment 66, whereinthe sexual dyad is a heterosexual dyad.

Specific Embodiment 68. The method of Specific Embodiment 66 or 67,wherein both members of the sexual dyad are of about 18 to about 50years and are trying to conceive.

The various embodiments described above can be combined to providefurther embodiments. All of the U.S. patents, U.S. patent applicationpublications, U.S. patent applications, foreign patents, foreign patentapplications and non-patent publications referred to in thisspecification and/or listed in the Application Data Sheet areincorporated herein by reference, in their entirety. Aspects of theembodiments can be modified, if necessary to employ concepts of thevarious patents, applications and publications to provide yet furtherembodiments.

These and other changes can be made to the embodiments in light of theabove-detailed description. In general, in the following claims, theterms used should not be construed to limit the claims to the specificembodiments disclosed in the specification and the claims, but should beconstrued to include all possible embodiments along with the full scopeof equivalents to which such claims are entitled. Accordingly, theclaims are not limited by the disclosure.

What is claimed is:
 1. A topical, isotonic composition comprising: (a) aprebiotic oligosaccharide; (b) a metal co-factor; and (c) borneol,pharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing (e.g., ester prodrugs such as bornyl acetate); wherein the pHof the composition ranges from about 3.5 to about
 8. 2. The topical,isotonic composition of claim 1, wherein the prebiotic oligosaccharidecomprises raffinose, lactulose, trehalose, gal actooligosaccharide,alpha-glucan oligosaccharide, beta-glucan oligosaccharide,fructooligosaccharide, isomaltooligosaccharide, maltose, inulin, pectin,or any combination thereof
 3. The topical, isotonic composition of claim1, wherein the metal co-factor comprises zinc, selenium, manganese,cobalt, iron, copper, or any combination thereof.
 4. The topical,isotonic composition of claim 1, wherein the composition comprisesbornyl acetate in an amount ranging from about 0.00025% to about 0.15%by weight of the composition.
 5. The topical, isotonic composition ofclaim 4, wherein the composition comprises from about 0.005% to about0.5% of an essential oil comprising said bornyl acetate by weight of theessential oil.
 6. The topical composition according to claim 1, whereinthe essential oil comprising bornyl acetate comprises essential oil fromJuniperus communis, Juniperus occidentalis, Juniperus scopulorum, Abiessibirica, Abies alba, Abies balsamea, Abies fraseri, Abies grandis,Abies spectabilis, Abies koreana, Abies procera, Abies nordmanniana,Abies magnifica, Abies pinsapo, Abies lasiocarpa, Abies concolor,Pseudotsuga menziesii, Rosmarinus officinalis or any combinationthereof.
 7. The topical, isotonic composition of claim 1, furthercomprising a sesquiterpene alcohol or monocylic sesquiterpene in anamount ranging from about 0.001% to about 0.1% by weight of thecomposition.
 8. The topical, isotonic composition of claim 1, furthercomprising a biofilm inhibiting agent in an amount ranging from about0.001% to about 0.16% by weight of the composition.
 9. The topical,isotonic composition of claim 1, further comprising a viscosityincreasing agent in an amount ranging from about 0.1% to about 10% byweight of the composition.
 10. The topical, isotonic composition ofclaim 1, wherein said composition is isotonic with physiological fluidsand/or skin and/or mucosa of the urogenital and/or anogenital region.11. The topical, isotonic composition of claim 10, wherein saidphysiological genital fluid is selected from cervico-vaginal fluids,urethral secretion, semen, or smegma.
 12. The topical, isotoniccomposition of claim 1, further comprising a preservative in an amountof about 0.001% to about 4% by weight of the composition.
 13. Thetopical, isotonic composition of claim 1, wherein the pH of thecomposition is about 4.5 or about 6.8.
 14. A method of hydrating theurogenital and/or anogenital region of a subject comprising topicallyadministering an effective amount of the topical, isotonic compositionof claim 1 to the urogenital and/or anogenital region of the subject.15. A method of lubricating the urogenital and/or anogenital region of asubject comprising topically administering an effective amount of thetopical, isotonic composition of claim 1 to the urogenital and/oranogenital region of the subject.
 16. A method of cleaning theurogenital and/or anogenital region of a subject comprising topicallyadministering an effective amount of the topical, isotonic compositionof claim 1 to the urogenital and/or anogenital region of the subject.17. A method of decreasing irritation or inflammation of the urogenitaland/or anogenital region of a subject comprising topically administeringan effective amount of the topical, isotonic composition of claim 1 tothe urogenital and/or anogenital region of the subject.
 18. A method ofhydrating, conditioning, cleansing or lubricating the urogenital and/oranogenital region of both members of a sexual dyad, wherein the sexualdyad comprises at least one female subject, comprising: (a)administering an effective amount of a first topical, isotoniccomposition to the urogenital and/or anogenital region of either or bothmembers of the sexual dyad during the non-fertile phase of the femalesubject, wherein the first topical, isotonic composition administeredduring the non-fertile phase and the first topical, isotonic compositionhas a pH of about 4.5 and a tonicity of about 150 mOsm/kg. (b)administering an effective amount of a second topical, isotoniccomposition to the urogenital and/or anogenital region of either or bothmembers of the sexual dyad during the fertile phase of the femalesubject, wherein the topical, isotonic composition administered duringthe fertile phase has a pH of about 6.8 and a tonicity of about 240mOsm/kg.
 19. The method according to claim 18, wherein said first andsecond topical, isotonic compositions are independently selected fromtopical, isotonic compositions comprising: a) a prebioticoligosaccharide; b) a metal co-factor; and c) borneol, orpharmaceutically acceptable salts thereof, or prodrugs of any of theforegoing (e.g., bornyl acetate).
 20. A method of hydrating,conditioning, cleansing or lubricating the urogenital and/or anogenitalregion of both members of a sexual dyad, wherein the sexual dyadcomprises at least one female subject, comprising: administering aneffective amount of a topical, isotonic composition to the urogenitaland/or anogenital region of either or both members of the sexual dyadduring the fertile phase of the female subject, wherein the topical,isotonic composition, includes a muco-adhesive, a biofilm inhibitor, ananti-inflammatory compound and a flavonoid antioxidant, administeredduring the fertile phase has a pH of about 5-6.8 and a tonicity of about240 mOsm/kg.
 21. The method according to claim 20, wherein said topical,isotonic composition comprises: a) a prebiotic oligosaccharide; b) ametal co-factor; c) borneol, or pharmaceutically acceptable saltsthereof, or prodrugs of any of the foregoing (e.g., bornyl acetate). 23.The method according to claim 22, wherein said topical, isotoniccomposition further comprises: d) a sesquiterpene alcohol (e.g., nerolioil); e) a biofilm inhibiting agent (e.g., a Lamiaceae essential oilsuch as Mentha spicate); f) a preservative of Lactobacillus ferment; andg) sodium dehydroacetate.